Classifications

• • C—CHEMISTRY; METALLURGY
  • C07—ORGANIC CHEMISTRY
  • C07D—HETEROCYCLIC COMPOUNDS
  • C07D401/00—Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, at least one ring being a six-membered ring with only one nitrogen atom
  • C07D401/02—Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, at least one ring being a six-membered ring with only one nitrogen atom containing two hetero rings
  • C07D401/04—Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, at least one ring being a six-membered ring with only one nitrogen atom containing two hetero rings directly linked by a ring-member-to-ring-member bond
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
  • A61P35/00—Antineoplastic agents
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
  • A61P43/00—Drugs for specific purposes, not provided for in groups A61P1/00-A61P41/00
• • C—CHEMISTRY; METALLURGY
  • C07—ORGANIC CHEMISTRY
  • C07D—HETEROCYCLIC COMPOUNDS
  • C07D401/00—Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, at least one ring being a six-membered ring with only one nitrogen atom
  • C07D401/14—Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, at least one ring being a six-membered ring with only one nitrogen atom containing three or more hetero rings
• • C—CHEMISTRY; METALLURGY
  • C07—ORGANIC CHEMISTRY
  • C07D—HETEROCYCLIC COMPOUNDS
  • C07D403/00—Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, not provided for by group C07D401/00
  • C07D403/02—Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, not provided for by group C07D401/00 containing two hetero rings
  • C07D403/04—Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, not provided for by group C07D401/00 containing two hetero rings directly linked by a ring-member-to-ring-member bond
• • C—CHEMISTRY; METALLURGY
  • C07—ORGANIC CHEMISTRY
  • C07D—HETEROCYCLIC COMPOUNDS
  • C07D403/00—Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, not provided for by group C07D401/00
  • C07D403/14—Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, not provided for by group C07D401/00 containing three or more hetero rings
• • C—CHEMISTRY; METALLURGY
  • C07—ORGANIC CHEMISTRY
  • C07D—HETEROCYCLIC COMPOUNDS
  • C07D405/00—Heterocyclic compounds containing both one or more hetero rings having oxygen atoms as the only ring hetero atoms, and one or more rings having nitrogen as the only ring hetero atom
  • C07D405/14—Heterocyclic compounds containing both one or more hetero rings having oxygen atoms as the only ring hetero atoms, and one or more rings having nitrogen as the only ring hetero atom containing three or more hetero rings

Definitions

• the present invention is directed to compounds, and salts and solvates thereof, their synthesis, and their use as modulators or inhibitors of the Raf enzyme.
• the compounds of the present invention are useful for modulating (e.g. inhibiting) Raf activity and for treating diseases or conditions mediated by Raf, such as for example, disease states associated with abnormal cell growth such as cancer.
• Erk pathway is an intracellular signal transduction pathway used by nearly all types of human cells to translate extracellular signals to cellular decisions, including proliferation, differentiation, senescence, or apoptosis (Wellbrock et al., Nat. Rev. MoI. Cell Biol. 11 :875-885 (2004)).
• Ras GTPase which receives signals from membrane receptors and activates the Raf protein kinases, which activate the Mek protein kinases, which in turn activate the Erk protein kinases.
• Activated Erk kinases phosphorylate a number of nuclear and cytoplasmic targets to initiate various cellular decisions.
• Raf The biological importance of Raf in the Erk pathway is underscored by the finding that mutated forms of Raf are associated with certain human malignancies (see e.g. Monia et al., Nature Medicine 2:668-675 (1996); Davies et al., Nature 417:949-954 (2002)).
• Three distinct genes have been identified in mammals that encode Raf proteins; a-Raf, b-Raf and c-Raf (also known as RaM) and isoformic variants that result from differential splicing of mRNA are known (Chong et al., EMBO J. 20:3716-3727 (2001)).
• the Erk pathway is mutationally activated in a number of human cancers, most often by mutation of the Ras or b-Raf genes. Mutations in Ras and b- Raf genes generally occur in the same tumor types, including cancers of the colon, lung and pancreas and melanoma, but are usually mutually exclusive. This suggests that activation of either Ras or Raf is sufficient for pathway activation and cancer progression.
• R 1 is H, -OH, (C 1 to C 6 ) alkyl, (C 2 to C 8 ) alkenyl, (C 2 to C 8 ) alkynyl, cyano, -N(R 5a R 5b ), -C(O)N(R 5a R 5b ), (C 3 to C 8 ) cycloalkyl, (C 6 to C 14 ) aryl, (C 2 to C 9 ) cycloheteroalkyl, or (C 2 to C 9 ) heteroaryl, wherein each of said (C 1 to C 6 ) alkyl, (C 2 to C 8 ) alkenyl, (C 2 to C 8 ) alkynyl, (C 3 to C 8 ) cycloalkyl, (C 6 to C i4 ) aryl, (C 2 to Cg) cycloheteroalkyl, and (C 2 to C 9 ) heteroaryl is optionally substituted with at least one
• R 3 is H, or-NH(CH 2 ) n R 7 ;
• R 4 is -SR 8 , -OR 8 , H, -OH, (C 1 to C 6 ) alkyl, (C 2 to C 8 ) alkenyl, (C 2 to C 8 ) alkynyl, cyano, -N(R 5a R 5b ), -C(O)N(R 5a R 5b ), (C 3 to C 8 ) cycloalkyl, (C 6 to C 14 ) aryl, (C 2 to C 9 ) cycloheteroalkyl, or (C 2 to C 9 ) heteroaryl, wherein each of said (C 3 to C 8 ) cycloalkyl, (C 6 to C 14 ) aryl, (C 2 to C 9 ) cycloheteroalkyl, and (C 2 to C 9 ) heteroaryl is optionally substituted with at least one R 6 group;
• R 5a and R 5b are each independently H, (C 1 to C 6 ) alkyl, (C 2 to C 8 ) alkenyl, (C 2 to C 8 ) alkynyl, (C 2 to C 9 ) cycloheteroalkyl, or (C 3 to C 8 ) cycloalkyl, wherein each of said (C 2 to C 9 ) cycloheteroalkyl and (C 3 to C 8 ) cycloalkyl is optionally substituted with at least one R 6 group;
• R 6 is -OH, (C 1 to C 6 ) alkyl, (C 2 to C 8 ) alkenyl, (C 2 to C 8 ) alkynyl, cyano, (C 3 to C 8 ) cycloalkyl, (C 2 to C 9 ) cycloheteroalkyl, (C 6 to C 14 ) aryl, (C 2 to C 9 ) heteroaryl, -(CH 2 ) n C
• R 7 is H, (C 1 to C 6 ) alkyl, (C 2 to C 8 ) alkenyl, (C 2 to C 8 ) alkynyl, -N(R 5a R 5b ), (C 2 to C 9 ) cycloheteroalkyl, (C 2 to C 9 ) heteroaryl, (C 3 to C 8 ) cycloalkyl, or (C 6 to C 14 ) aryl, wherein each of said (C 1 to C 6 ) alkyl, (C 2 to C 8 ) alkenyl, (C 2 to C 8 ) alkynyl, (C 2 to C 9 ) cycloheteroalkyl, (C 3 to C 8 ) cycloalkyl, (C 6 to C 14 ) aryl, and (C 2 to Cg) heteroaryl are optionally substituted with at least one group selected from halogen, -OH, (C 1 to C 6 ) alkyl, -N(R 5a R
• R 8 is (C 1 to C 6 ) alkyl, (C 2 to C 8 ) alkenyl, (C 2 to C 8 ) alkynyl, -(CH 2 ) n -cyano, -(CH 2 ) n -(C 3 to C 8 ) cycloalkyl, -(CH 2 V(C 6 to C 14 ) aryl, -(CH 2 J n -(C 2 to C 9 ) cycloheteroalkyl, or -(CH 2 ) ⁇ -(C 2 to C 9 ) heteroaryl;
• R 9 is H, -OH, (C 1 to C 6 ) alkyl, (C 2 to C 8 ) alkenyl, (C 2 to C 8 ) alkynyl, (C 1 to C 8 ) alkoxy, or
• R 1Oa and R 10b are each independently H, (C 1 to C 6 ) alkyl, (C 2 to C 8 ) alkenyl, (C 2 to C 8 ) alkynyl, (C 2 to C 9 ) cycloheteroalkyl, or (C 3 to C 8 ) cycloalkyl;
• X is N or CH; m is 0, 1 , 2, or 3; each n is independently 0, 1 , 2, 3, or 4; p is 0, 1 , 2, 3, or 4; or a salt or solvate thereof.
• R 7 is H, (C 1 to C 6 ) alkyl, (C 2 to C 8 ) alkenyl, (C 2 to C 8 ) alkynyl, -N(R 5a R 5b ), (C 2 to C 9 ) cycloheteroalkyl, or (C 3 to C 8 ) cycloalkyl, wherein each of said (C 1 to C 6 ) alkyl, (C 2 to C 8 ) alkenyl, (C 2 to C 8 ) alkynyl, (C 2 to C 9 ) cycloheteroalkyl, and (C 3 to C 8 ) cycloalkyl are optionally substituted with at least one group selected from
• p is 1 or 2 and R 1 is cyano or (C 1 to C 6 ) alkyl.
• p is 0 and R 1 is H or (C 1 to C 6 ) alkyl. In a still further aspect p is 0 or 1 and R 1 is (C 2 to C 9 ) cycloheteroalkyl and is optionally substituted with at least one R 6 group. In a still further aspect, the (C 2 to C 9 ) cycloheteroalkyl is selected from the group consisting of:
• each of said (C 2 to C 9 ) cycloheteralkyl groups are optionally substituted with at least one R 6 group.
• each R 2 is independently halogen, -OCH 3 , or (C 1 to C 6 ) alkyl.
• R 2 is independently halogen, -OCH 3 , C 1 alkyl, C 2 alkyl, C 3 alkyl, or C 4 alkyl.
• m is 1 and the compound of Formula (I) has the following structure:
• R 2 is Cl or CH 3 .
• R 7 is H, (C 1 to C 6 ) alkyl, (C 2 to C 9 ) cycloheteroalkyl, or (C 3 to C 8 ) cycloalkyl, wherein each of said (Ci to C 6 ) alkyl, (C 2 to C 9 ) cycloheteroalkyl, - A -
• (C 3 to C 8 ) cycloalkyl are optionally substituted with at least one group selected from halogen, -OH, (C 1 to C 6 ) alkyl, -N(R 5a R 5b ), (C 3 to C 8 ) cycloalkyl, (C 6 to C 14 ) aryl, and (C 2 to C 9 ) cycloheteroalkyl.
• R 3 is -NH(CH 2 ) n R 7 where n is 1 or 2, and R 7 is (C-i to C 6 ) alkyl and is substituted with one -OH group.
• R 7 is (C-i to C 6 ) alkyl and is substituted with one -OH group.
• R 3 is a compound of Formula (I), as described above, wherein X is N and R 4 is H, (C 2 to C 9 ) cycloheteroalkyl, -OR 8 , or -N(R 5a R 5b ), wherein said (C 2 to C 9 ) cycloheteroalkyl is optionally substituted with at least one R 6 group.
• the present invention is a compound of Formula (I)
• R 1 is H, -OH, (C 1 to C 6 ) alkyl, (C 2 to C 8 ) alkenyl, (C 2 to C 8 ) alkynyl, cyano, -N(R 5a R 5b ),
• each R 2 is independently H, halogen, (C 1 to C 6 ) alkyl, (C 2 to C 8 ) alkenyl, (C 2 to C 8 ) alkynyl, (C 1 to C 8 ) alkoxy, or cyano;
• R 3 is H, or -NR 11 (CH 2 ) n R 7 ;
• R 4 is -SR 8 , -OR 8 , H, -OH, (C 1 to C 6 ) alkyl, (C 2 to C 8 ) alkenyl, (C 2 to C 8 ) alkynyl, cyano, -N(R 5a R 5b ), -C(O)N(R 5a R 5b ), (C 3 to C 8 ) cycloalkyl, (C 6 to C i4 ) aryl, (C 2 to C 9 ) cycloheteroalkyl, or (C 2 to C 9 ) heteroaryl, wherein each of said (C 3 to C 8 ) cycloalkyl, (C 6 to C 14 ) aryl, (C 2 to C 9 ) cycloheteroalkyl, and (C 2 to Cg) heteroaryl is optionally substituted with at least one R 6 group;
• R 5a and R 5b are each independently H, (C 1 to C 6 ) alkyl, (C 2 to C 8 ) alkenyl, (C 2 to C 8 ) alkynyl, (C 2 to C 9 ) cycloheteroalkyl, or (C 3 to C 8 ) cycloalkyl, wherein each of said (C 2 to C 9 ) cycloheteroalkyl and (C 3 to C 8 ) cycloalkyl is optionally substituted with at least one R 6 group;
• R s is -OH, (C 1 to C 6 ) alkyl, (C 2 to C 8 ) alkenyl, (C 2 to C 8 ) alkynyl, cyano, (C 3 to C 8 ) cycloalkyl, (C 2 to C 9 ) cycloheteroalkyl, (C 6 to C 14 ) aryl, (C 2 to C 9 ) heteroaryl, -(CH 2 ) n C(O)R 9 , or -N(R 10a R 10b );
• R 7 is H, (C 1 to C 6 ) alkyl, (C 2 to C 8 ) alkenyl, (C 2 to C 8 ) alkynyl, -N(R 5a R 5b ), (C 2 to C 9 ) cycloheteroalkyl, (C 2 to C 9 ) heteroaryl, (C 3 to C 8 ) cycloalkyl, or (C 6 to C 14 ) aryl, wherein each of said (C-i to C 6 ) alkyl, (C 2 to C 8 ) alkenyl, (C 2 to C 8 ) alkynyl, (C 2 to C 9 ) cycloheteroalkyl, (C 3 to C 8 ) cycloalkyl, (C 6 to Cu) aryl, and (C 2 to C 9 ) heteroaryl are optionally substituted with at least one group selected from halogen, -OH, (C 1 to C 6 ) alkyl, -N(R 5a R
• R 9 is H, -OH, (C 1 to C 6 ) alkyl, (C 2 to C 8 ) alkenyl, (C 2 to C 8 ) alkynyl, (C 1 to C 8 ) alkoxy, or
• R 1Oa and R 1Ob are each independently H, (C 1 to C 6 ) alkyl, (C 2 to C 8 ) alkenyl, (C 2 to C 8 ) alkynyl, (C 2 to C 9 ) cycloheteroalkyl, or (C 3 to C 8 ) cycloalkyl;
• R 11 is H, (C 1 to C 6 ) alkyl, (C 2 to C 8 ) alkenyl, (C 2 to C 8 ) alkynyl, (C 2 to C 9 ) cycloheteroalkyl, (C 2 to C 9 ) heteroaryl, (C 3 to C 8 ) cycloalkyl, or (C 6 to Ci 4 ) aryl, wherein each of said (C 1 to C 6 ) alkyl, (C 2 to C 8 ) alkenyl, (C 2 to C 8 ) alkynyl, (C 2 to C 9 ) cycloheteroalkyl, (C 3 to C 8 ) cycloalkyl, (C 6 to C 14 ) aryl, and (C 2 to C 9 ) heteroaryl are optionally substituted with at least one group selected from halogen, -OH, (C-i to C 6 ) alkyl, -N(R 5a R 5b ), (C 3 to C 8
• R 12 is (C 1 to C 6 ) alkyl, (C 2 to C 8 ) alkenyl, (C 2 to C 8 ) alkynyl, -N(R 5a R 5b ), (C 2 to C 9 ) cycloheteroalkyl, (C 2 to C 9 ) heteroaryl, (C 3 to C 8 ) cycloalkyl, or (C 6 to C 14 ) aryl, wherein each of said (C 1 to C 6 ) alkyl, (C 2 to C 8 ) alkenyl, (C 2 to C 8 ) alkynyl, (C 2 to C 9 ) cycloheteroalkyl, (C 3 to C 8 ) cycloalkyl, (C 6 to C 14 ) aryl, and (C 2 to C 9 ) heteroaryl are optionally substituted with at least one group selected from halogen, -OH, (C 1 to C 6 ) alkyl, -N(R 5a R 5
• R 7 is H, (C 1 to C 6 ) alkyl, (C 2 to C 8 ) alkenyl, (C 2 to C 8 ) alkynyl, -N(R 5a R 5b ), (C 2 to C 9 ) cycloheteroalkyl, (C 2 to C 9 ) heteroaryl, (C 3 to C 8 ) cycloalkyl, or (C 6 to C 14 ) aryl, wherein each of said (C 1 to C 6 ) alkyl, (C 2 to C 8 ) alkenyl, (C 2 to C 8 ) alkynyl, (C 2 to C 9 ) cycloheteroalkyl, (C 3 to C 8 ) cycloalkyl, (C 6 to C 14 ) aryl, and (C 2 to C 9 ) heteroaryl are optionally substituted with at least one group selected from halogen, -OH,
• R 7 is H, (C 1 to C 6 ) alkyl, (C 2 to C 9 ) cycloheteroalkyl, or (C 3 to C 8 ) cycloalkyl, wherein each of said (C 1 to C 6 ) alkyl, (C 2 to C 9 ) cycloheteroalkyl, and (C 3 to C 8 ) cycloalkyl are optionally substituted with at least one group selected from halogen, -OH, (C 1 to C 6 ) alkyl, -N(R 5a R 5b ), -OC(O)R 12 , (C 3 to C 8 ) cycloalkyl, (C 6 to C 14 ) aryl, and (C 2 to C 9 ) cycloheteroalkyl.
• R 7 is H, (C 1 to C 6 ) alkyl, (C 2 to C 9 ) cycloheteroalkyl, or (C 3 to C 8 ) cycloalkyl,
• R 3 is -NR 11 (CH 2 ) n R 7 .
• R 7 is H, (C 1 to C 6 ) alkyl, (C 2 to C 9 ) cycloheteroalkyl, or (C 3 to C 8 ) cycloalkyl, wherein each of said (C-i to C 6 ) alkyl, (C 2 to C 9 ) cycloheteroalkyl, and (C 3 to C 8 ) cycloalkyl are optionally substituted with at least one group selected from halogen, -OH, (C 1 to C 6 ) alkyl, -N(R 5a R 5b ), -OC(O)R 12 , (C 3 to C 8 ) cycloalkyl, (C 6 to C 14 ) aryl, and (C 2 to C 9 ) cycloheteroalkyl.
• R 3 is -NH(CH 2 ) n R 7 where n is
• a further aspect of the present invention is a compound according to Formula (I), as described above, which is selected from the group consisting of: 3-chloro-5-(1-isopropyl-4-pyrimidin-4-yl-1 H-pyrazol- 3-yl)phenol; 3-[4-(2-anilinopyrimidin-4-yl)-1-methyl-1 H-pyrazol-3-yl]-5-chlorophenol; 2-(3-(3-chloro-5- hydroxyphenyl)-4-(2-((S)-2-hydroxypropylamino)pyrimidin-4-yl)-1W-pyrazol-1-yl)acetonitrile; 2-(3-(3-
• the compound is selected from the group consisting of 3-chloro-5-(1-isopropyl-4-pyrimidin-4- yl-1 H-pyrazol-3-yl)phenol; 3-[4-(2-anilinopyrimidin-4-yl)-1-methyl-1 H-pyrazol-3-yl]-5-chlorophenol; 2-(3-(3- chloro-5-hydroxyphenyl)-4-(2-((S)-2-hydroxypropylamino)pyrimidin-4-yl)-1H-pyrazol-1-yl)acetonitrile; 2-(3- (3-Chloro-5-hydroxyphenyl-4-(2-(2-hydroxyethylamino)pyrimidin-4-yl)-1H-pyrazol-1-yl)acetonitrile; and 3- [3-(3-hydroxy-5-methylphenyl)-4-(2- ⁇ [(2S)-2-hydroxypropyl]amino ⁇ pyrimidin-4-yl)-1 H-pyrazol
• the compound is selected from the group consisting of 3-(3-(3- hydroxy-5-methylphenyl)-4-(2-((R)-2-hydroxypropylamino)pyrimidin-4-yl)-1H-pyrazol-1-yl)propanenitrile; 3- chloro-5-[4-[2-(isobutylamino)pyrimidin-4-yl]-1-(1-methylpiperidin-4-yl)-1 H-pyrazol-3-yl]phenol; 3-chloro-5- ⁇ 4-[2-(isobutylamino)pyrimidin-4-yl]-1-piperidin-4-yl-1H-pyrazol-3-yl ⁇ phenol; 3-chloro-5-[4- ⁇ 2-[(2-hydroxy-2- methylpropyl)amino]pyrimidin-4-yl ⁇ -1-(1-methylpiperidin-4-yl)-1W-pyrazol-3-yl]phenol; and 3-chloro-5- ⁇ 4-[2- (iso)
• the compound is selected from the group consisting of 3-chloro-5-[4- ⁇ 2-[(2-hydroxy-2-methylpropyl)amino]pyrimidin-4-yl ⁇ -1- (1-methylpiperidin-4-yl)-1/-/-pyrazol-3-yl]phenol; 3-chloro-5- ⁇ 4-[2-(isobutylamino)pyrimidin-4-yl]-1-methyl- 1H-pyrazol-3-yl ⁇ phenol; ⁇ 3-(3-chloro-5-hydroxyphenyl)-4-[2-(tetrahydrofuran-3-ylamino)pyrimidin-4-yl]-1 H- pyrazol-1-yl ⁇ acetonitrile; [3-(3-chloro-5-hydroxyphenyl)-4- ⁇ 2-[(2-hydroxy-1-methylethyl)amino]pyrimidin-4- yl ⁇ -1H-pyrazol-1-yl]acetonitrile; and [4-(2-amino
• the compound is selected from the group consisting of [3-(3- chloro-5-hydroxyphenyl)-4-(2- ⁇ [(2S)-2-hydroxypropyl]amino ⁇ pyrimidin-4-yl)-1 H-pyrazol-1-yl]acetonitrile; 3- [4- ⁇ 2-[(2-hydroxyethyl)amino]pyrimidin-4-yl ⁇ -3-(3-hydroxy-5-methylphenyl)-1H-pyrazol-1-yl]propanenitrile; 3- ⁇ 3-(3-hydroxy-5-methylphenyl)-4-[2-(tetrahydrofuran-3-ylamino)pyrimidin-4-yl]-1 H-pyrazol-1- yl ⁇ propanenitrile; [3-(3-chloro-5-hydroxyphenyl)-4-(2- ⁇ [(1S)-2-hydroxy-1-methylethyl]amino ⁇ pyrimidin-4-yl)- 1 H-pyrazol-1-yl]acetonitrile
• a further aspect of the present invention is a compound according to Formula (I), as described above, which is selected from the group consisting of 2-(3-(3-fluoro-5-hydroxyphenyl)-4-(2-((S)-2- hydroxypropylamino)pyrimidin-4-yl)-1H-pyrazol-1-yl)acetonitrile, 2-(3-(3-hydroxy-5-methylphenyl)-4-(2- ((S)-2-hydroxypropylamino)pyrimidin-4-yl)-1 H-pyrazol-1-yl)-acetonitrile, 3-(1-(cyanomethyl)-4-(2-(2- hydroxyethylamino)pyrimidin-4-yl)-1 H-pyrazol-3-yl)-5-hydroxybenzonitrile, 3-(4-(2-((S)-2- hydroxypropylamino)pyrimidin-4-yl)-1-((5-methylisoxazol-3-yl)methyl)-1 H-pyrazol-3-
• the present invention also relates to a pharmaceutical composition, comprising at least one compound as described herein, or a pharmaceutically acceptable salt thereof, and a pharmaceutically acceptable carrier or diluent.
• the present invention also relates to a method of treating abnormal cell growth, or any Raf- mediated disease or condition, in a mammal in need thereof, comprising the step of administering to said mammal a therapeutically effective amount of at least one compound as described herein, or a pharmaceutically acceptable salt thereof.
• the abnormal cell growth is cancerous.
• the abnormal cell growth in non-cancerous.
• the present invention further relates to a method of inhibiting Raf enzymatic activity, comprising contacting a Raf enzyme with a Raf-inhibiting amount of at least one compound as described herein, or a pharmaceutically acceptable salt thereof.
• the present invention further relates to the use of any of the compounds as described herein, or a salt or solvate thereof, in the manufacture of a medicament for the treatment of abnormal cell growth in a mammal.
• the present invention further relates to methods of making the compounds as described herein using the methods as shown in the specific examples herein and in the general synthetic methods A, B, C, D, and E, as described herein.
• the present invention further relates to any of the compounds described above, or salts or solvates thereof, for use as a medicament.
• the present invention further relates to the use of any of the compounds described above, or salts or solvates thereof, for the manufacture of a medicament for the treatment of abnormal cell growth.
• halo and/or halogen refer to fluorine, chlorine, bromine or iodine.
• (C 1 to Ce) alkyl refers to a saturated aliphatic hydrocarbon radical including straight chain and branched chain groups of 1 to 6 carbon atoms.
• Examples of (C 1 to C 6 ) alkyl groups include methyl, ethyl, propyl, 2-propyl, ⁇ -butyl, /so-butyl, ferf-butyl, pentyl, and the like.
• (C 2 to C 8 ) alkenyl means an alkyl moiety comprising 2 to 8 carbons having at least one carbon-carbon double bond.
• the carbon-carbon double bond in such a group may be anywhere along the 2 to 8 carbon chain that will result in a stable compound.
• Such groups include both the E and Z isomers of said alkenyl moiety. Examples of such groups include, but are not limited to, ethenyl, propenyl, butenyl, allyl, and pentenyl.
• (C 2 to C 8 ) alkynyl means an alkyl moiety comprising from 2 to 8 carbon atoms and having at least one carbon-carbon triple bond.
• the carbon-carbon triple bond in such a group may be anywhere along the 2 to 8 carbon chain that will result in a stable compound. Examples of such groups include, but are not limited to, ethyne, propyne, 1-butyne, 2-butyne, 1-pentyne, 2-pentyne, 1- hexyne, 2-hexyne, and 3-hexyne.
• (C 1 to C 8 ) alkoxy means an O-alkyl group wherein said alkyl group contains from 1 to 8 carbon atoms and is straight, branched, or cyclic.
• alkyl group contains from 1 to 8 carbon atoms and is straight, branched, or cyclic.
• examples of such groups include, but are not limited to, methoxy, ethoxy, n-propyloxy, iso-propyloxy, n-butoxy, iso-butoxy, tert-butoxy, cyclopentyloxy, and cyclohexyloxy.
• (C 1 to C 8 ) heteroalkyl refers to a straight- or branched-chain alkyl group having a total of from 2 to 12 atoms in the chain, including from 1 to 8 carbon atoms, and one or more atoms of which is a heteroatom selected from S, O, and N, with the proviso that said chain may not contain two adjacent O atoms or two adjacent S atoms.
• the S atoms in said chains may be optionally oxidized with one or two oxygen atoms, to afford sulfides and sulfones, respectively.
• the (C-i to C 8 ) heteroalkyl groups in the compounds of the present invention can contain an oxo group at any carbon or heteroatom that will result in a stable compound.
• exemplary (C 1 to C 8 ) heteroalkyl groups include, but are not limited to, alcohols, alkyl ethers, primary, secondary, and tertiary alkyl amines, amides, ketones, esters, sulfides, and sulfones.
• (C 6 to C 14 ) aryl means a group derived from an aromatic hydrocarbon containing from 6 to 14 carbon atoms. Examples of such groups include, but are not limited to, phenyl or naphthyl.
• (C 2 to C 9 ) heteroaryl means an aromatic heterocyclic group having a total of from 5 to 10 atoms in its ring, and containing from 2 to 9 carbon atoms and from one to four heteroatoms each independently selected from O, S and N, and with the proviso that the ring of said group does not contain two adjacent O atoms or two adjacent S atoms.
• the heterocyclic groups include benzo-fused ring systems.
• aromatic heterocyclic groups are pyridinyl, imidazolyl, pyrimidinyl, pyrazolyl, triazolyl, pyrazinyl, tetrazolyl, furyl, thienyl, isoxazolyl, thiazolyl, oxazolyl, isothiazolyl, pyrrolyl, quinolinyl, isoquinolinyl, indolyl, benzimidazolyl, benzofuranyl, cinnolinyl, indazolyl, indolizinyl, phthalazinyl, pyridazinyl, triazinyl, isoindolyl, pteridinyl, purinyl, oxadiazolyl, thiadiazolyl, furazanyl, benzofurazanyl, benzothiophenyl, benzothiazolyl, benzoxazolyl, quinazolinyl, quinox
• the C 2 to C 9 heteroaryl groups may be C-attached or N-attached where such is possible.
• a group derived from pyrrole may be pyrrol-1-yl (N-attached) or pyrrol-3-yl (C-attached).
• a group derived from imidazole may be imidazol-1-yl (N-attached) or imidazol-3-yl (C-attached).
• (C 2 to C 9 ) cycloheteroalkyl means a non-aromatic, monocyclic, bicyclic, tricyclic, spirocyclic, or tetracyclic group having a total of from 4 to 13 atoms in its ring system, and containing from 2 to 9 carbon atoms and from 1 to 4 heteroatoms each independently selected from O, S and N, and with the proviso that the ring of said group does not contain two adjacent O atoms or two adjacent S atoms.
• such C 2 to C 9 cycloheteroalkyl groups may contain an oxo substituent at any available atom that will result in a stable compound.
• such a group may contain an oxo atom at an available carbon or nitrogen atom. Such a group may contain more than one oxo substituent if chemically feasible.
• a C 2 to C 9 cycloheteroalkyl group contains a sulfur atom, said sulfur atom may be oxidized with one or two oxygen atoms to afford either a sulfoxide or sulfone.
• An example of a 4 membered cycloheteroalkyl group is azetidinyl (derived from azetidine).
• An example of a 5 membered cycloheteroalkyl group is pyrrolidinyl.
• An example of a 6 membered cycloheteroalkyl group is piperidinyl.
• An example of a 9 membered cycloheteroalkyl group is indolinyl.
• An example of a 10 membered cycloheteroalkyl group is 4H-quinolizinyl.
• C 2 to Cg cycloheteroalkyl groups include, but are not limited to, tetrahydrofuranyl, dihydrofuranyl, tetrahydrothienyl, tetrahydropyranyl, dihydropyranyl, tetrahydrothiopyranyl, piperidino, morpholino, thiomorpholino, thioxanyl, piperazinyl, azetidinyl, oxetanyl, thietanyl, homopiperidinyl, oxepanyl, thiepanyl, oxazepinyl, diazepinyl, thiazepinyl, 1 ,2,3,6-tetrahydropyridinyl, 2-pyrrolinyl, 3-pyrrolinyl, indolinyl, 2H-pyranyl, 4H- pyranyl, dioxanyl, 1 ,3-dioxolany
• (C 3 to C 8 ) cycloalkyl group means a saturated, monocyclic, fused, spirocyclic, or polycyclic ring structure having a total of from 3 to 8 carbon ring atoms.
• groups include, but are not limited to, cyclopropyl, cyclobutyl, cyclopentyl, cyclopentenyl, cyclohexyl, cycloheptyl, and adamantyl.
• cyano refers to a -C ⁇ N group.
• substituted means that the specified group or moiety bears one or more substituents.
• unsubstituted means that the specified group bears no substituents.
• optionally substituted means that the specified group is unsubstituted or substituted by one or more substituents. It is to be understood that in the compounds of the present invention when a group is said to be “unsubstituted,” or is “substituted” with fewer groups than would fill the valencies of all the atoms in the compound, the remaining valencies on such a group are filled by hydrogen.
• a C 6 aryl group also called “phenyl” herein
• phenyl is substituted with one additional substituent
• one of ordinary skill in the art would understand that such a group has 4 open positions left on carbon atoms of the C 6 aryl ring (6 initial positions, minus one to which the remainder of the compound of the present invention is bonded, minus an additional substituent, to leave 4).
• the remaining 4 carbon atoms are each bound to one hydrogen atom to fill their valencies.
• a C 6 aryl group in the present compounds is said to be "disubstituted," one of ordinary skill in the art would understand it to mean that the C 6 aryl has 3 carbon atoms remaining that are unsubstituted.
• solvate is used to describe a molecular complex between compounds of the present invention and solvent molecules.
• solvates include, but are not limited to, compounds of the invention in combination water, isopropanol, ethanol, methanol, dimethylsulfoxide (DMSO), ethyl acetate, acetic acid, ethanolamine, or mixtures thereof.
• DMSO dimethylsulfoxide
• hydrate can be used when said solvent is water. It is specifically contemplated that in the present invention one solvent molecule can be associated with one molecule of the compounds of the present invention, such as a hydrate.
• solvates of the present invention are contemplated as solvates of compounds of the present invention that retain the biological effectiveness of the non-hydrate form of the compounds.
• pharmaceutically acceptable salt means a salt of a compound of the present invention that retains the biological effectiveness of the free acids and bases of the specified derivative and that is not biologically or otherwise undesirable.
• pharmaceutically acceptable formulation means a combination of a compound of the invention, or a salt or solvate thereof, and a carrier, diluent, and/or excipient(s) that are compatible with a compound of the present invention, and is not deleterious to the recipient thereof.
• Pharmaceutical formulations can be prepared by procedures known to those of ordinary skill in the art.
• the compounds of the present invention can be formulated with common excipients, diluents, or carriers, and formed into tablets, capsules, and the like.
• excipients, diluents, and carriers that are suitable for such formulations include the following: fillers and extenders such as starch, sugars, mannitol, and silicic derivatives; binding agents such as carboxymethyl cellulose and other cellulose derivatives, alginates, gelatin, and polyvinyl pyrrolidone; moisturizing agents such as glycerol; disintegrating agents such as povidone, sodium starch glycolate, sodium carboxymethylcellulose, agar, calcium carbonate, and sodium bicarbonate; agents for retarding dissolution such as paraffin; resorption accelerators such as quaternary ammonium compounds; surface active agents such as cetyl alcohol, glycerol monostearate; adsorptive carriers such as kaolin and bentonite; and lubricants such as talc, calcium and magnesium
• Final pharmaceutical forms may be pills, tablets, powders, lozenges, saches, cachets, or sterile packaged powders, and the like, depending on the type of excipient used. Additionally, it is specifically contemplated that pharmaceutically acceptable formulations of the present invention can contain more than one active ingredient. For example, such formulations may contain more than one compound according to the present invention. Alternatively, such formulations may contain one or more compounds of the present invention and one or more additional agents that reduce abnormal cell growth.
• the term "Raf-inhibiting amount” as used herein, refers to the amount of a compound of the present invention, or a salt or solvate thereof, required to inhibit the enzymatic activity of Raf in vivo, such as in a mammal, or in vitro.
• inhibiting Raf enzyme activity means decreasing the activity or functioning of the Raf enzyme either in vitro or in vivo, such as in a mammal, such as a human, by contacting the enzyme with a compound of the present invention.
• Raf as used herein means a-Raf, b-Raf, c-Raf, or mutants thereof, or any of the known Raf isoformic splice variants.
• therapeutically effective amount means an amount of a compound of the present invention, or a salt or solvate thereof, that, when administered to a mammal in need of such treatment, is sufficient to effect treatment, as defined herein.
• a therapeutically effective amount of a compound of the present invention, or a salt or solvate thereof is a quantity sufficient to modulate or inhibit the activity of the Raf enzyme such that a disease condition that is mediated by activity of the Raf enzyme is reduced or alleviated.
• treat with reference to abnormal cell growth, or to any Raf mediated disease or condition, in a mammal, particularly a human, include: (i) preventing the disease or condition from occurring in a subject which may be predisposed to the condition, such that the treatment constitutes prophylactic treatment for the pathologic condition; (ii) modulating or inhibiting the disease or condition, i.e., arresting its development; (iii) relieving the disease or condition, i.e., causing regression of the disease or condition; or (iv) relieving and/or alleviating the disease or condition or the symptoms resulting from the disease or condition, e.g., relieving an inflammatory response without addressing the underlying disease or condition.
• abnormal cell growth such as cancer
• these terms simply mean that the life expectancy of an individual affected with abnormal cell growth will be increased or that one or more of the symptoms of the disease will be reduced.
• references herein to the inventive compounds include references to salts, solvates, and complexes thereof, including polymorphs, stereoisomers, tautomers, and isotopically labeled versions thereof.
• compounds of the present invention can be pharmaceutically acceptable salts and/or pharmaceutically acceptable solvates.
• tumor cells tumor cells
• tumors that proliferate by expressing a mutated tyrosine kinase or overexpression of a receptor tyrosine kinase
• benign and malignant cells of other proliferative diseases in which aberrant tyrosine kinase activation occurs any tumors that proliferate by receptor tyrosine kinases; any tumors that proliferate by aberrant serine/threonine kinase activation; benign and malignant cells of other proliferative diseases in which aberrant serine/threonine kinase activation occurs
• tumors both benign and malignant, expressing an activated Ras oncogene
• tumor cells both benign and malignant, in which the Ras protein is activated as a result of oncogenic mutation in another gene
• benign and malignant cells of other proliferative diseases in which aberrant Ras activation occurs the abnormal growth of: tumor cells (tumors) that proliferate by expressing a mutated tyros
• abnormal cell growth also refers to and includes the abnormal growth of cells, both benign and malignant, resulting from activity of the enzyme farnesyl protein transferase.
• abnormal cell growth and “hyperproliferative disorder” are used interchangeably in this application.
• stereoisomers refers to compounds that have identical chemical constitution, but differ with regard to the arrangement of their atoms or groups in space.
• enantiomers refers to two stereoisomers of a compound that are non-superimposable mirror images of one another.
• racemic or “racemic mixture,” as used herein, refer to a 1:1 mixture of enantiomers of a particular compound.
• diastereomers refers to the relationship between a pair of stereoisomers that comprise two or more asymmetric centers and are not mirror images of one another.
• the compounds of the present invention are useful for modulating or inhibiting Raf activity.
• these compounds are useful for the prevention and/or treatment of disease states associated with abnormal cell growth such as cancer, alone or in combination with other anti-cancer agents.
• ⁇ is used in structural formulas herein to depict the bond that is the point of attachment of the moiety or substituent to the core or backbone structure.
• the carbon atoms and their bound hydrogen atoms are not explicitly depicted, e.g., ' ⁇ ⁇ represents a methyl
• the compounds of the present invention may have asymmetric carbon atoms.
• the carbon- carbon bonds of the compounds of the present invention may be depicted herein using a solid line ( ), a solid wedge ( ⁇ m ⁇ m ⁇ ), or a dotted wedge ( "'" ).
• the use of a solid line to depict bonds to asymmetric carbon atoms is meant to indicate that all possible stereoisomers (e.g. specific enantiomers, racemic mixtures, etc.) at that carbon atom are included.
• the use of either a solid or dotted wedge to depict bonds to asymmetric carbon atoms is meant to indicate that only the stereoisomer shown is meant to be included. It is possible that compounds of the invention may contain more than one asymmetric carbon atom.
• a solid line to depict bonds to asymmetric carbon atoms is meant to indicate that all possible stereoisomers are meant to be included.
• the compounds of the present invention can exist as enantiomers and diastereomers or as racemates and mixtures thereof.
• the use of a solid line to depict bonds to one or more asymmetric carbon atoms in a compound of the invention and the use of a solid or dotted wedge to depict bonds to other asymmetric carbon atoms in the same compound is meant to indicate that a mixture of diastereomers is present.
• enantiomers include chiral synthesis from a suitable optically pure precursor or resolution of the reacemate using, for example, chiral high pressure liquid chromatography (HPLC).
• HPLC high pressure liquid chromatography
• the racemate (or a racemic precursor) may be reacted with a suitable optically active compound, for example, an alcohol, or, in the case where the compound contains an acidic or basic moiety, an acid or base such as tartaric acid or 1-phenylethylamine.
• a suitable optically active compound for example, an alcohol, or, in the case where the compound contains an acidic or basic moiety, an acid or base such as tartaric acid or 1-phenylethylamine.
• the resulting diastereomeric mixture may be separated by chromatography and/or fractional crystallization and one or both of the diastereoisomers converted to the corresponding pure enantiomer(s) by means well known to one skilled in the art.
• Chiral compounds of the invention may be obtained in enantiomerically-enriched form using chromatography, typically HPLC, on an asymmetric resin with a mobile phase consisting of a hydrocarbon, typically heptane or hexane, containing from 0 to 50% isopropanol, typically from 2 to 20%, and from 0 to 5% of an alkylamine, typically 0.1% diethylamine. Concentration of the eluate affords the enriched mixture.
• Stereoisomeric conglomerates may be separated by conventional techniques known to those skilled in the art. See, e.g. "Stereochemistry of Organic Compounds" by E L Eliel (Wiley, New York, 1994), the disclosure of which is incorporated herein by reference in its entirety.
• a compound of the invention contains an alkenyl or alkenylene group
• geometric cisltrans (or Z/E) isomers are possible.
• the compound contains, for example, a keto or oxime group or an aromatic moiety
• tautomeric isomerism ('tautomerism') can occur.
• Examples of tautomerism include keto and enol tautomers.
• a single compound may exhibit more than one type of isomerism. Included within the scope of the invention are all stereoisomers, geometric isomers and tautomeric forms of the inventive compounds, including compounds exhibiting more than one type of isomerism, and mixtures of one or more thereof.
• Cis/trans isomers may be separated by conventional techniques well known to those skilled in the art, for example, chromatography and fractional crystallization.
• the compounds of the present invention may be administered as prodrugs.
• certain derivatives of compounds of Formula (I) which may have little or no pharmacological activity themselves can, when administered to a mammal, be converted into a compound of Formula (I) having the desired activity, for example, by hydrolytic cleavage.
• Such derivatives are referred to as "prodrugs”.
• Prodrugs can, for example, be produced by replacing appropriate functionalities present in the compound of Formula (I) with certain moieties known to those skilled in the art. See, e.g. "Pro-drugs as Novel Delivery Systems", Vol. 14, ACS Symposium Series (T Higuchi and W Stella) and "Bioreversible Carriers in Drug Design", Pergamon Press, 1987 (ed.
• prodrugs include: an ester moiety in the place of a carboxylic acid functional group; an ether moiety or an amide moiety in place of an alcohol functional group; and an amide moiety in place of a primary or secondary amino functional group.
• replacement groups are known to those of skill in the art. See, e.g. "Design of Prodrugs” by H Bundgaard (Elsevier, 1985), the disclosure of which is incorporated herein by reference in its entirety. It is also possible that certain compounds of Formula (I) may themselves act as prodrugs of other compounds of Formula (I).
• Salts of the present invention can be prepared according to methods known to those of skill in the art.
• Examples of salts include, but are not limited to, acetate, acrylate, benzenesulfonate, benzoate (such as chlorobenzoate, methylbenzoate, dinitrobenzoate, hydroxybenzoate, and methoxybenzoate), bicarbonate, bisulfate, bisulfite, bitartrate, borate, bromide, butyne-1 ,4-dioate, calcium edetate, camsylate, carbonate, chloride, caproate, caprylate, clavulanate, citrate, decanoate, dihydrochloride, dihydrogenphosphate, edetate, edislyate, estolate, esylate, ethylsuccinate, formate, fumarate, gluceptate, gluconate, glutamate, glycollate, glycollylarsanilate, heptanoate, he
• the compounds of the present invention that are basic in nature are capable of forming a wide variety of different salts with various inorganic and organic acids. Although such salts must be pharmaceutically acceptable for administration to animals, it is often desirable in practice to initially isolate the compound of the present invention from the reaction mixture as a pharmaceutically unacceptable salt and then simply convert the latter back to the free base compound by treatment with an alkaline reagent and subsequently convert the latter free base to a pharmaceutically acceptable acid addition salt.
• the acid addition salts of the base compounds of this invention can be prepared by treating the base compound with a substantially equivalent amount of the selected mineral or organic acid in an aqueous solvent medium or in a suitable organic solvent, such as methanol or ethanol. Upon evaporation of the solvent, the desired solid salt is obtained.
• the desired acid salt can also be precipitated from a solution of the free base in an organic solvent by adding an appropriate mineral or organic acid to the solution.
• Those compounds of the present invention that are acidic in nature are capable of forming base salts with various pharmacologically acceptable cations.
• such salts include the alkali metal or alkaline-earth metal salts and particularly, the sodium and potassium salts. These salts are all prepared by conventional techniques.
• the chemical bases which are used as reagents to prepare the pharmaceutically acceptable base salts of this invention are those which form non-toxic base salts with the acidic compounds of the present invention.
• Such non-toxic base salts include those derived from such pharmacologically acceptable cations as sodium, potassium calcium and magnesium, etc.
• salts can be prepared by treating the corresponding acidic compounds with an aqueous solution containing the desired pharmacologically acceptable cations, and then evaporating the resulting solution to dryness, preferably under reduced pressure.
• they may also be prepared by mixing lower alkanolic solutions of the acidic compounds and the desired alkali metal alkoxide together, and then evaporating the resulting solution to dryness in the same manner as before.
• stoichiometric quantities of reagents are preferably employed in order to ensure completeness of reaction and maximum yields of the desired final product.
• the desired salt may be prepared by any suitable method available in the art, for example, treatment of the free base with an inorganic acid, such as hydrochloric acid, hydrobromic acid, sulfuric acid, nitric acid, phosphoric acid and the like, or with an organic acid, such as acetic acid, maleic acid, succinic acid, mandelic acid, fumaric acid, malonic acid, pyruvic acid, oxalic acid, glycolic acid, salicylic acid, a pyranosidyl acid, such as glucuronic acid or galacturonic acid, an alpha-hydroxy acid, such as citric acid or tartaric acid, an amino acid, such as aspartic acid or glutamic acid, an aromatic acid, such as benzoic acid or cinnamic acid, a sulfonic acid, such as p-toluenesulfonic acid or ethanesulfonic acid, or the like.
• an inorganic acid such as hydrochloric acid, hydrobro
• the desired salt may be prepared by any suitable method, for example, treatment of the free acid with an inorganic or organic base, such as an amine (primary, secondary or tertiary), an alkali metal hydroxide or alkaline earth metal hydroxide, or the like.
• an inorganic or organic base such as an amine (primary, secondary or tertiary), an alkali metal hydroxide or alkaline earth metal hydroxide, or the like.
• suitable salts include organic salts derived from amino acids, such as glycine and arginine, ammonia, primary, secondary, and tertiary amines, and cyclic amines, such as piperidine, morpholine and piperazine, and inorganic salts derived from sodium, calcium, potassium, magnesium, manganese, iron, copper, zinc, aluminum and lithium.
• inventive compounds, agents and salts may exist in different crystal or polymorphic forms, all of which are intended to be within the scope of the present invention and specified formulas.
• the invention also includes isotopically-labeled compounds of the invention, wherein one or more atoms is replaced by an atom having the same atomic number, but an atomic mass or mass number different from the atomic mass or mass number usually found in nature.
• isotopes suitable for inclusion in the compounds of the invention include isotopes of hydrogen, such as 2 H and 3 H, carbon, such as 11 C, 13 C and 14 C, chlorine, such as 36 CI, fluorine, such as 18 F, iodine, such as 123 I and 125 I, nitrogen, such as 13 N and 15 N, oxygen, such as 15 O, 17 O and 18 O, phosphorus, such as 32 P, and sulfur, such as 35 S.
• isotopically-labeled compounds of the invention for example, those incorporating a radioactive isotope, are useful in drug and/or substrate tissue distribution studies.
• radioactive isotopes tritium, 3 H, and carbon-14, 14 C are particularly useful for this purpose in view of their ease of incorporation and ready means of detection. Substitution with heavier isotopes such as deuterium, 2 H, may afford certain therapeutic advantages resulting from greater metabolic stability, for example, increased in vivo half-life or reduced dosage requirements, and hence may be preferred in some circumstances. Substitution with positron emitting isotopes, such as 11 C, 18 F, 15 O and 13 N, can be useful in Positron Emission Topography (PET) studies for examining substrate receptor occupancy.
• PET Positron Emission Topography
• Isotopically-labeled compounds of the invention can generally be prepared by conventional techniques known to those skilled in the art or by processes analogous to those described herein, using an appropriate isotopically-labeled reagent in place of the non-labeled reagent otherwise employed.
• compositions of the invention comprise a therapeutically effective amount of at least one compound of the present invention and an inert, pharmaceutically acceptable carrier or diluent.
• a pharmaceutical composition of the invention is administered in a suitable formulation prepared by combining a therapeutically effective amount (i.e., a Raf modulating, regulating, or inhibiting amount effective to achieve therapeutic efficacy) of at least one compound of the present invention (as an active ingredient) with one or more pharmaceutically suitable carriers, which may be selected, for example, from diluents, excipients and auxiliaries that facilitate processing of the active compounds into the final pharmaceutical preparations.
• a therapeutically effective amount i.e., a Raf modulating, regulating, or inhibiting amount effective to achieve therapeutic efficacy
• one compound of the present invention as an active ingredient
• pharmaceutically suitable carriers which may be selected, for example, from diluents, excipients and auxiliaries that facilitate processing of the active compounds into the final pharmaceutical preparations.
• the pharmaceutical carriers employed may be either solid or liquid.
• Exemplary solid carriers are lactose, sucrose, talc, gelatin, agar, pectin, acacia, magnesium stearate, stearic acid and the like.
• Exemplary liquid carriers are syrup, peanut oil, olive oil, water and the like.
• the inventive compositions may include time-delay or time-release material known in the art, such as glyceryl monostearate or glyceryl distearate alone or with a wax, ethylcellulose, hydroxypropylmethylcellulose, methylmethacrylate or the like. Further additives or excipients may be added to achieve the desired formulation properties.
• a bioavailability enhancer such as Labrasol, Gelucire or the like, or formulator, such as CMC (carboxy-methylcellulose), PG (propyleneglycol), or PEG (polyethyleneglycol), may be added.
• CMC carboxy-methylcellulose
• PG propyleneglycol
• PEG polyethyleneglycol
• Gelucire ® a semi-solid vehicle that protects active ingredients from light, moisture and oxidation, may be added, e.g., when preparing a capsule formulation.
• the preparation can be tableted, placed in a hard gelatin capsule in powder or pellet form, or formed into a troche or lozenge.
• the amount of solid carrier may vary, but generally will be from about 25 mg to about 1 g.
• the preparation may be in the form of syrup, emulsion, soft gelatin capsule, sterile injectable solution or suspension in an ampoule or vial or non-aqueous liquid suspension.
• a semi-solid carrier is used, the preparation may be in the form of hard and soft gelatin capsule formulations.
• the inventive compositions are prepared in unit-dosage form appropriate for the mode of administration, e.g. parenteral or oral administration.
• a salt of a compound of the present invention may be dissolved in an aqueous solution of an organic or inorganic acid, such as a 0.3 M solution of succinic acid or citric acid.
• the agent may be dissolved in a suitable co-solvent or combinations of co-solvents.
• suitable co-solvents include alcohol, propylene glycol, polyethylene glycol 300, polysorbate 80, glycerin and the like in concentrations ranging from 0 to 60% of the total volume.
• a compound of the present invention is dissolved in DMSO and diluted with water.
• the composition may also be in the form of a solution of a salt form of the active ingredient in an appropriate aqueous vehicle such as water or isotonic saline or dextrose solution.
• the agents of the compounds of the present invention may be formulated into aqueous solutions, preferably in physiologically compatible buffers such as Hanks solution, Ringer's solution, or physiological saline buffer.
• physiologically compatible buffers such as Hanks solution, Ringer's solution, or physiological saline buffer.
• penetrants appropriate to the barrier to be permeated are used in the formulation. Such penetrants are generally known in the art.
• the compounds can be formulated by combining the active compounds with pharmaceutically acceptable carriers known in the art.
• Such carriers enable the compounds of the invention to be formulated as tablets, pills, dragees, capsules, liquids, gels, syrups, slurries, suspensions and the like, for oral ingestion by a subject to be treated.
• Pharmaceutical preparations for oral use can be obtained using a solid excipient in admixture with the active ingredient (agent), optionally grinding the resulting mixture, and processing the mixture of granules after adding suitable auxiliaries, if desired, to obtain tablets or dragee cores.
• Suitable excipients include: fillers such as sugars, including lactose, sucrose, mannitol, or sorbitol; and cellulose preparations, for example, maize starch, wheat starch, rice starch, potato starch, gelatin, gum, methyl cellulose, hydroxypropylmethyl-cellulose, sodium carboxymethylcellulose, or polyvinylpyrrolidone (PVP).
• PVP polyvinylpyrrolidone
• disintegrating agents may be added, such as crosslinked polyvinyl pyrrolidone, agar, or alginic acid or a salt thereof such as sodium alginate.
• Dragee cores are provided with suitable coatings.
• suitable coatings For this purpose, concentrated sugar solutions may be used, which may optionally contain gum arabic, polyvinyl pyrrolidone, Carbopol gel, polyethylene glycol, and/or titanium dioxide, lacquer solutions, and suitable organic solvents or solvent mixtures.
• Dyestuffs or pigments may be added to the tablets or dragee coatings for identification or to characterize different combinations of active agents.
• compositions that can be used orally include push-fit capsules made of gelatin, as well as soft, sealed capsules made of gelatin and a plasticizer, such as glycerol or sorbitol.
• the push-fit capsules can contain the active ingredients in admixture with fillers such as lactose, binders such as starches, and/or lubricants such as talc or magnesium stearate, and, optionally, stabilizers.
• the active agents may be dissolved or suspended in suitable liquids, such as fatty oils, liquid paraffin, or liquid polyethylene glycols.
• stabilizers may be added. All formulations for oral administration should be in dosages suitable for such administration.
• the compositions may take the form of tablets or lozenges formulated in conventional manner.
• the compounds for use according to the present invention may be conveniently delivered in the form of an aerosol spray presentation from pressurized packs or a nebuliser, with the use of a suitable propellant, e.g., dichlorodifluoromethane, trichlorofluoromethane, dichlorotetrafluoroethane, carbon dioxide or other suitable gas.
• a suitable propellant e.g., dichlorodifluoromethane, trichlorofluoromethane, dichlorotetrafluoroethane, carbon dioxide or other suitable gas.
• a suitable propellant e.g., dichlorodifluoromethane, trichlorofluoromethane, dichlorotetrafluoroethane, carbon dioxide or other suitable gas.
• a suitable propellant e.g., dichlorodifluoromethane, trichlorofluoromethane, dichlorotetrafluoroethane, carbon dioxide
• the compounds may be formulated for parenteral administration by injection, e.g., by bolus injection or continuous infusion.
• Formulations for injection may be presented in unit-dosage form, e.g., in ampoules or in multi-dose containers, with an added preservative.
• the compositions may take such forms as suspensions, solutions or emulsions in oily or aqueous vehicles, and may contain formulatory agents such as suspending, stabilizing and/or dispersing agents.
• compositions for parenteral administration include aqueous solutions of the active compounds in water-soluble form. Additionally, suspensions of the active agents may be prepared as appropriate oily injection suspensions. Suitable lipophilic solvents or vehicles include fatty oils such as sesame oil, or synthetic fatty acid esters, such as ethyl oleate or triglycerides, or liposomes. Aqueous injection suspensions may contain substances that increase the viscosity of the suspension, such as sodium carboxymethyl cellulose, sorbitol, or dextran. Optionally, the suspension may also contain suitable stabilizers or agents that increase the solubility of the compounds to allow for the preparation of highly concentrated solutions.
• the active ingredient may be in powder form for constitution with a suitable vehicle, e.g. sterile pyrogen-free water, before use.
• a suitable vehicle e.g. sterile pyrogen-free water
• the compounds of the present invention may also be formulated as a depot preparation.
• Such long-acting formulations may be administered by implantation (for example, subcutaneously or intramuscularly) or by intramuscular injection.
• the compounds may be formulated with suitable polymeric or hydrophobic materials (for example, as an emulsion in an acceptable oil) or ion-exchange resins, or as sparingly soluble derivatives, for example, as a sparingly soluble salt.
• a pharmaceutical carrier for hydrophobic compounds is a co- solvent system comprising benzyl alcohol, a non-polar surfactant, a water-miscible organic polymer, and an aqueous phase.
• the co-solvent system may be a VPD co-solvent system.
• VPD is a solution of 3% w/v benzyl alcohol, 8% w/v of the non-polar surfactant polysorbate 80, and 65% w/v polyethylene glycol 300, made up to volume in absolute ethanol.
• the VPD co-solvent system (VPD: 5W) contains VPD diluted 1 :1 with a 5% dextrose in water solution. This co-solvent system dissolves hydrophobic compounds well, and itself produces low toxicity upon systemic administration.
• the proportions of a co- solvent system may be suitably varied without destroying its solubility and toxicity characteristics.
• co-solvent components may be varied: for example, other low-toxicity non- polar surfactants may be used instead of polysorbate 80; the fraction size of polyethylene glycol may be varied; other biocompatible polymers may replace polyethylene glycol, e.g. polyvinyl pyrrolidone; and other sugars or polysaccharides may be substituted for dextrose.
• other delivery systems for hydrophobic pharmaceutical compounds may be employed. Liposomes and emulsions are known examples of delivery vehicles or carriers for hydrophobic drugs. Certain organic solvents such as dimethylsulfoxide also may be employed, although usually at the cost of greater toxicity due to the toxic nature of DMSO.
• the compounds may be delivered using a sustained-release system, such as semipermeable matrices of solid hydrophobic polymers containing the therapeutic agent.
• sustained-release materials have been established and are known by those skilled in the art.
• Sustained-release capsules may, depending on their chemical nature, release the compounds for a few weeks up to over 100 days.
• additional strategies for protein stabilization may be employed.
• the pharmaceutical compositions also may comprise suitable solid- or gel-phase carriers or excipients. These carriers and excipients may provide marked improvement in the bioavailability of poorly soluble drugs.
• Such carriers or excipients include calcium carbonate, calcium phosphate, sugars, starches, cellulose derivatives, gelatin, and polymers such as polyethylene glycols. Furthermore, additives or excipients such as Gelucire®, Capryol®, Labrafil®, Labrasol®, Lauroglycol®, Plurol®, Peceol® Transcutol® and the like may be used.
• the pharmaceutical composition may be incorporated into a skin patch for delivery of the drug directly onto the skin.
• an exemplary daily dose generally employed will be from about 0.001 to about 1000 mg/kg of body weight, with courses of treatment repeated at appropriate intervals.
• the pharmaceutically acceptable formulations of the present invention may contain a compound of the present invention, or a salt or solvate thereof, in an amount of about 10 mg to about 2000 mg, or from about 10 mg to about 1500 mg, or from about 10 mg to about 1000 mg, or from about 10 mg to about 750 mg, or from about 10 mg to about 500 mg, or from about 25 mg to about 500 mg, or from about 50 to about 500 mg, or from about 100 mg to about 500mg.
• the pharmaceutically acceptable formulations of the present invention may contain a compound of the present invention, or a salt or solvate thereof, in an amount from about 0.5 w/w% to about 95 w/w%, or from about 1 w/w% to about 95 w/w%, or from about 1 w/w% to about 75 w/w%, or from about 5 w/w% to about 75 w/w%, or from about 10 w/w% to about 75 w/w%, or from about 10 w/w% to about 50 w/w%.
• the compounds of the present invention may be administered to a mammal suffering from abnormal cell growth, such as a human, either alone or as part of a pharmaceutically acceptable formulation, once a day, twice a day, three times a day, or four times a day, or even more frequently.
• This invention also relates to a method for the treatment of abnormal cell growth in a mammal, including a human, comprising administering to said mammal an amount of a compound of the Formula (I), as defined above, or a salt or solvate thereof, that is effective in treating abnormal cell growth.
• the abnormal cell growth is cancer, including, but not limited to, mesothelioma, hepatobiliary (hepatic and billiary duct), a primary or secondary CNS tumor, a primary or secondary brain tumor, lung cancer (NSCLC and SCLC), bone cancer, pancreatic cancer, skin cancer, cancer of the head or neck, cutaneous or intraocular melanoma, ovarian cancer, colon cancer, rectal cancer, cancer of the anal region, stomach cancer, gastrointestinal (gastric, colorectal, and duodenal), breast cancer, uterine cancer, carcinoma of the fallopian tubes, carcinoma of the endometrium, carcinoma of the cervix, carcinoma of the vagina, carcinoma of the vulva, Hodgkin's Disease, cancer of the esophagus, cancer of the small intestine, cancer of the endocrine system, cancer of the thyroid gland, cancer of the parathyroid gland, cancer of the adrenal gland, sarcoma of soft tissue, cancer of
• the cancer is selected from lung cancer (NSCLC and SCLC), cancer of the head or neck, ovarian cancer, colon cancer, rectal cancer, cancer of the anal region, stomach cancer, breast cancer, cancer of the kidney or ureter, renal cell carcinoma, carcinoma of the renal pelvis, neoplasms of the central nervous system (CNS), primary CNS lymphoma, non hodgkins's lymphoma, spinal axis tumors, or a combination of one or more of the foregoing cancers.
• lung cancer NSCLC and SCLC
• SCLC central nervous system
• the cancer is selected from lung cancer (NSCLC and SCLC), ovarian cancer, colon cancer, rectal cancer, cancer of the anal region, or a combination of one or more of the foregoing cancers.
• the cancer is selected from lung cancer (NSCLC and SCLC), ovarian cancer, colon cancer, rectal cancer, or a combination of one or more of the foregoing cancers.
• said abnormal cell growth is a benign proliferative disease, including, but not limited to, psoriasis, benign prostatic hypertrophy or restinosis.
• This invention also relates to a method for the treatment of abnormal cell growth in a mammal which comprises administering to said mammal an amount of a compound of the present invention, or a salt or solvate thereof, that is effective in treating abnormal cell growth in combination with an anti-tumor agent selected from the group consisting of mitotic inhibitors, alkylating agents, anti-metabolites, intercalating antibiotics, growth factor inhibitors, cell cycle inhibitors, enzymes, topoisomerase inhibitors, biological response modifiers, antibodies, cytotoxics, anti-hormones, and anti-androgens.
• an anti-tumor agent selected from the group consisting of mitotic inhibitors, alkylating agents, anti-metabolites, intercalating antibiotics, growth factor inhibitors, cell cycle inhibitors, enzymes, topoisomerase inhibitors, biological response modifiers, antibodies, cytotoxics, anti-hormones, and anti-androgens.
• the anti-tumor agent used in conjunction with a compound of the present invention and pharmaceutical compositions described herein is an anti- angiogenesis agent, kinase inhibitor, pan kinase inhibitor or growth factor inhibitor.
• Preferred pan kinase inhibitors include SutentTM (sunitinib), described in U.S. Patent No. 6,573,293 (Pfizer, Inc, NY, USA).
• Anti-angiogenesis agents include but are not limited to the following agents, such as EGF inhibitors, EGFR inhibitors, VEGF inhibitors, .
• VEGFR inhibitors TIE2 inhibitors, IGF1 R inhibitors, COX-II (cyclooxygenase II) inhibitors, MMP-2 (matrix-metalloprotienase 2) inhibitors, and MMP-9 (matrix- metalloprotienase 9) inhibitors.
• VEGF inhibitors include for example, Avastin (bevacizumab), an anti-VEGF monoclonal antibody of Genentech, Inc. of South San Francisco, California. Additional VEGF inhibitors include CP-547,632 (Pfizer Inc., NY, USA), AG13736 (Pfizer Inc.), ZD-6474 (AstraZeneca), AEE788
• IM862 Cytran Inc. of Kirkland, Washington, USA
• angiozyme a synthetic ribozyme from Ribozyme (Boulder, Colorado) and Chiron (Emeryville, California) and combinations thereof.
• VEGF inhibitors useful in the practice of the present invention are described in US Patent No. 6,534,524 and 6,235,764, both of which are incorporated in their entirety for all purposes. Additional VEGF inhibitors are described in, for example in WO 99/24440, in WO 95/21613, WO 99/61422, U.S. Patent 5,834,504, WO 98/50356, U.S. Patent 5,883,113 U.S. Patent 5,886,020, U.S. Patent 5,792,783, U.S.
• anti-angiogenic compounds include acitretin, fenretinide, thalidomide, zoledronic acid, angiostatin, aplidine, cilengtide, combretastatin A-4, endostatin, halofuginone, rebimastat, removab, Revlimid, squalamine, ukrain, Vitaxin and combinations thereof.
• Other antiproliferative agents that may be used in combination with the compounds of the present invention include inhibitors of the enzyme farnesyl protein transferase and inhibitors of the receptor tyrosine kinase PDGFr, including the compounds disclosed and claimed in the following: U.S. Patent 6,080,769; U.S.
• Each of the foregoing patents and patent applications is herein incorporated by reference in their entirety.
• PDGRr inhibitors include but are not limited to those disclosed in international patent application publication numbers WO01/40217 and WO2004/020431 , the contents of which are incorporated in their entirety for all purposes.
• Preferred PDGFr inhibitors include Pfizer's CP-673,451 and CP-868,596 and its salts.
• Preferred GARF inhibitors include Pfizer's AG-2037 (pelitrexol and its salts). GARF inhibitors useful in the practice of the present invention are disclosed in US Patent No. 5,608,082 which is incorporated in its entirety for all purposes.
• COX-II inhibitors which can be used in conjunction with a compound of Formula (I) and pharmaceutical compositions disclosed herein include CELEBREXTM (celecoxib), parecoxib, deracoxib, ABT-963, MK-663 (etoricoxib), COX-189 (Lumiracoxib), BMS 347070, RS 57067, NS-398, Bextra (valdecoxib), paracoxib, Vioxx (rofecoxib), SD-8381 , 4-Methyl-2-(3,4-dimethylphenyl)-1- (4-sulfamoyl-phenyl)-1 H-pyrrole, 2-(4-Ethoxyphenyl)-4-methyl-1-(4-sulfamoylphenyl)-1 H-pyrrole, T-614, JTE-522, S-2474, SVT-2016, CT-3, SC-58125 and Arcoxia (etoricoxib).
• COX-II CELEBREXTM
• the anti-tumor agent is celecoxib (U.S. Patent No. 5,466,823), valdecoxib (U.S. Patent No. 5,633,272), parecoxib (U.S. Patent No. 5,932,598), deracoxib (U.S. Patent No. 5,521,207), SD-8381 (U.S. Patent No. 6,034,256, Example 175), ABT-963 (WO 2002/24719), rofecoxib (CAS No. 162011-90-7), MK-663 (or etoricoxib) as disclosed in WO 1998/03484, COX-189 (Lumiracoxib) as disclosed in WO 1999/11605, BMS-347070 (U.S.
• Patent 6,180,651 NS-398 (CAS 123653-11-2), RS 57067 (CAS 17932-91-3), 4-Methyl-2-(3,4-dimethylphenyl)-1-(4-sulfamoyl-phenyl)-1 H- pyrrole, 2-(4-Ethoxyphenyl)-4-methyl-1-(4-sulfamoylphenyl)-1H-pyrrole, or meloxicam.
• NSAIDs non-steroidal antiinflammatory drugs
• NSAIDs non-steroidal antiinflammatory drugs
• NSAIDs non-steroidal antiinflammatory drugs
• NSAIDs non-steroidal antiinflammatory drugs
• NSAIDs non-steroidal antiinflammatory drugs
• NSAIDs non-steroidal antiinflammatory drugs
• NSAIDs non-steroidal antiinflammatory drugs
• NSAIDs non-steroidal antiinflammatory drugs
• NSAIDs non-steroidal antiinflammatory drugs
• Preferred COX-I inhibitors include ibuprofen (Motrin), nuprin, naproxen (Aleve), indomethacin (Indocin), nabumetone (Relafen) and combinations thereof.
• Targeted agents used in combination with a compound of the present invention and pharmaceutical compositions disclosed herein include EGFr inhibitors such as lressa (gefitinib, AstraZeneca), Tarceva (erlotinib or OSI-774, OSI Pharmaceuticals Inc.), Erbitux (cetuximab, lmclone Pharmaceuticals, Inc.), EMD-7200 (Merck AG), ABX-EGF (Amgen Inc.
• EGFr inhibitors include lressa, Erbitux, Tarceva and combinations thereof.
• anti-tumor agents include those selected from pan erb receptor inhibitors or ErbB2 receptor inhibitors, such as CP-724,714 (Pfizer, Inc.), CI-1033 (canertinib, Pfizer, Inc.), Herceptin (trastuzumab, Genentech Inc.), Omitarg (2C4, pertuzumab, Genentech Inc.), TAK-165 (Takeda), GW-572016 (lonafarnib, GlaxoSmithKline), GW-282974 (GlaxoSmithKline), EKB-569 (Wyeth), PKM 66 (Novartis), dHER2 (HER2 Vaccine, Corixa and GlaxoSmithKline), APC8024 (HER2 Vaccine, Dendreon), anti- HER2/neu bispecific antibody (Decof Cancer Center), B7.her2.lgG3 (Agensys), AS HER2 (Research Institute for Rad Biology & Medicine), trifunctional bispecific antibodies (Univers
• pan erbb receptor inhibitors include GW572016, CI-1033, EKB-569, and Omitarg and combinations thereof.
• Additional erbB2 inhibitors include those disclosed in WO 98/02434, WO 99/35146, WO 99/35132, WO 98/02437, WO 97/13760, WO 95/19970, U.S. Patent 5,587,458, and U.S. Patent 5,877,305, each of which is herein incorporated by reference in its entirety.
• ErbB2 receptor inhibitors useful in the present invention are also disclosed in U.S. Patents 6,465,449, and 6,284,764, and in WO 2001/98277 each of which are herein incorporated by reference in their entirety.
• anti-tumor agents may be selected from the following agents, BAY-43-9006 (Onyx Pharmaceuticals Inc.), Genasense (augmerosen, Genta), Panitumumab (Abgenix/Amgen), Zevalin (Schering), Bexxar (Corixa/GlaxoSmithKline), Abareiix, Alimta, EPO 906 (Novartis), discodermolide (XAA- 296), ABT-510 (Abbott), Neovastat (Aeterna), enzastaurin (EIi Lilly), Combrestatin A4P (Oxigene), ZD-6126 (AstraZeneca), flavopiridol (Aventis), CYC-202 (Cyclacei), AVE-8062 (Aventis), DMXAA (Roche/Antisoma), Thymitaq (Eximias), Temodar (temozolomide, Schering Plough) and Revilimd (Celegene) and combinations
• Histerelin histrelin acetate
• Plenaixis abareiix depot
• Atrasentan ABT-627
• Satraplatin JM-216
• thalomid Thalidomide
• Theratope Temilifene (DPPE), ABI-007 (paclitaxel), Evista (raloxifene), Atamestane (Biomed-777), Xyotax (polyglutamate paclitaxel), Targetin (bexarotine) and combinations thereof.
• anti-tumor agents may be selected from the following agents, Trizaone (tirapazamine), Aposyn (exisulind), Nevastat (AE-941), Ceplene (histamine dihydrochloride), Orathecin (rubitecan), Virulizin, Gastrimmune (G17DT), DX-8951f (exatecan mesylate), Onconase (ranpirnase), BEC2 (mitumoab), Xcytrin (motexafin gadolinium) and combinations thereof.
• anti-tumor agents may be selected from the following agents, CeaVac (CEA), NeuTrexin
• Additional anti-tumor agents may be selected from the following agents, OvaRex (oregovomab), Osidem (IDM-1), and combinations thereof. Additional anti-tumor agents may be selected from the following agents, Advexin (ING 201), Tirazone (tirapazamine), and combinations thereof. Additional anti-tumor agents may be selected from the following agents, RSR13
• Additional anti-tumor agents may be selected from the following agents, Canvaxin, GMK vaccine, PEG lnteron A, Taxoprexin (DHA/paciltaxel), and combinations thereof.
• anti-tumor agents include Pfizer's MEK1/2 inhibitor PD325901, Array Biopharm's MEK inhibitor ARRY-142886, Bristol Myers' CDK2 inhibitor BMS-387,032, Pfizer's CDK inhibitor PD0332991 and AstraZeneca's AXD-5438, and combinations thereof.
• mTOR inhibitors may also be utilized such as CCI-779 (Wyeth) and rapamycin derivatives RAD001 (Novartis) and AP-23573 (Ariad), HDAC inhibitors, SAHA (Merck Inc./Aton Pharmaceuticals) and combinations thereof.
• Additional anti-tumor agents include aurora 2 inhibitor VX-680 (Vertex), and Chk1/2 inhibitor XL844 (Exilixis).
• cytotoxic agents e.g., one or more selected from the group consisting of epirubicin (Ellence), docetaxel (Taxotere), paclitaxel, Zinecard (dexrazoxane), rituximab (Rituxan) imatinib mesylate (Gleevec), and combinations thereof, may be used in combination with a compound of the present invention and pharmaceutical compositions disclosed herein.
• the invention also contemplates the use of the compounds of the present invention together with hormonal therapy, including but not limited to, exemestane (Aromasin, Pfizer Inc.), leuprorelin (Lupron or Leuplin, TAP/Abbott/Takeda), anastrozole (Arimidex, Astrazeneca), gosrelin (Zoladex, AstraZeneca), doxercalciferol, fadrozole, formestane, tamoxifen citrate (tamoxifen, Nolvadex, AstraZeneca), Casodex (AstraZeneca), Abarelix (Praecis), Trelstar, and combinations thereof.
• exemestane Amasin, Pfizer Inc.
• leuprorelin Louprorelin
• anastrozole Arimidex, Astrazeneca
• gosrelin Zoladex, AstraZeneca
• doxercalciferol
• the invention also relates to the use of the compounds of the present invention together with hormonal therapy agents such as anti-estrogens including, but not limited to fulvestrant, toremifene, raloxifene, lasofoxifene, letrozole (Femara, Novartis), anti-androgens such as bicalutamide, flutamide, mifepristone, nilutamide, CasodexTM(4'-cyano-3-(4-fluorophenylsulphonyl)-2-hydroxy-2-methyl-3'-
• hormonal therapy agents such as anti-estrogens including, but not limited to fulvestrant, toremifene, raloxifene, lasofoxifene, letrozole (Femara, Novartis), anti-androgens such as bicalutamide, flutamide, mifepristone, nilutamide, CasodexTM(4'-cyano-3-(
• the invention provides a compound of the present invention alone or in combination with one or more supportive care products, e.g., a product selected from the group consisting of Filgrastim (Neupogen), ondansetron (Zofran), Fragmin, Procrit, Aloxi, Emend, or combinations thereof.
• supportive care products e.g., a product selected from the group consisting of Filgrastim (Neupogen), ondansetron (Zofran), Fragmin, Procrit, Aloxi, Emend, or combinations thereof.
• Particularly preferred cytotoxic agents include Camptosar, Erbitux, Iressa, Gleevec, Taxotere and combinations thereof.
• topoisomerase I inhibitors may be utilized as anti-tumor agents: camptothecin; irinotecan HCI (Camptosar); edotecarin; orathecin (Supergen); exatecan (Daiichi); BN-80915 (Roche); and combinations thereof.
• Particularly preferred toposimerase Il inhibitors include epirubicin (Ellence).
• Alkylating agents include, but are not limited to, nitrogen mustard N-oxide, cyclophosphamide, ifosfamide, melphalan, busulfan, mitobronitoi, carboquone, thiotepa, ranimustine, nimustine, temozolomide, AMD-473, altretamine, AP-5280, apaziquone, brostallicin, bendamustine, carmustine, estramustine, fotemustine, glufosfamide, ifosfamide, KW-2170, mafosfamide, and mitolactol; platinum- coordinated alkylating compounds include but are not limited to, cisplatin, Paraplatin (carboplatin), eptaplatin, lobaplatin, nedaplatin, Eloxatin (oxaliplatin, Sanofi) or satrplatin and combinations thereof. Particularly preferred alkylating agents include Eloxatin (oxalip
• Antimetabolites include but are not limited to, methotrexate, 6-mercaptopurine riboside, mercaptopurine, 5-fluorouracil (5-FU) alone or in combination with leucovorin, tegafur, UFT, doxifluridine, carmofur, cytarabine, cytarabine ocfosfate, enocitabine, S-1 , Alimta (premetrexed disodium, LY231514,
• MTA gemzar
• Gemzar gemcitabine, EIi Lilly
• fludarabin 5-azacitidine, capecitabine, cladribine, clofarabine, decitabine, eflornithine, ethynylcytidine, cytosine arabinoside, hydroxyurea, TS-1 , melphalan, nelarabine, nolatrexed, ocfosfate, disodium premetrexed, pentostatin, pelitrexol, raltitrexed, triapine, trimetrexate, vidarabine, vincristine, vinorelbine; or for example, one of the preferred anti-metabolites disclosed in European Patent Application No.
• Antibiotics include intercalating antibiotics and include, but are not limited to: aclarubicin, actinomycin D, amrubicin, annamycin, adriamycin, bleomycin, daunorubicin, doxorubicin, elsamitrucin, epirubicin, galarubicin, idarubicin, mitomycin C, nemorubicin, neocarzinostatin, peplomycin, pirarubicin, rebeccamycin, stimalamer, streptozocin, valrubicin, zinostatin and combinations thereof.
• Plant derived anti-tumor substances include for example those selected from mitotic inhibitors, for example vinblastine, docetaxel (Taxotere), paclitaxel and combinations thereof.
• Cytotoxic topoisomerase inhibiting agents include one or more agents selected from the group consisting of aclarubicn, amonafide, belotecan, camptothecin, 10-hydroxycamptothecin, 9- aminocamptothecin, diflomotecan, irinotecan HCI (Camptosar), edotecarin, epirubicin (Ellence), etoposide, exatecan, gimatecan, lurtotecan, mitoxantrone, pirarubicin, pixantrone, rubitecan, sobuzoxane, SN-38, tafluposide, topotecan, and combinations thereof.
• Preferred cytotoxic topoisomerase inhibiting agents include one or more agents selected from the group consisting of camptothecin, 10-hydroxycamptothecin, 9-aminocamptothecin, irinotecan HCI (Camptosar), edotecarin, epirubicin (Ellence), etoposide, SN-38, topotecan, and combinations thereof.
• Interferons include interferon alpha, interferon alpha-2a, interferon, alpha-2b, interferon beta, interferon gamma-1a, interferon gamma-1b (Actimmune), or interferon gamma-n1 and combinations thereof.
• agents include filgrastim, lentinan, sizofilan, TheraCys, ubenimex, WF-10, aldesleukin, alemtuzumab, BAM-002, dacarbazine, daclizumab, denileukin, gemtuzumab ozogamicin, ibritumomab, imiquimod, lenograstim, lentinan, melanoma vaccine (Corixa), molgramostim, OncoVAX-CL, sargramostim, tasonermin, tecleukin, thymalasin, tositumomab, Virulizin, Z-100, epratuzumab, mitumomab, oregovomab, pemtumomab (Y- muHMFGI), Provenge (Dendreon) and combinations thereof.
• Biological response modifiers are agents that modify defense mechanisms of living organisms or biological responses, such as survival, growth, or differentiation of tissue cells to direct them to have antitumor activity.
• agents include krestin, lentinan, sizofiran, picibanil, ubenimex and combinations thereof.
• Velcade (bortemazib, Millenium), tretinoin, and combinations thereof.
• Platinum-coordinated compounds include but are not limited to, cisplatin, carboplatin, nedaplatin, oxaliplatin, and combinations thereof.
• Camptothecin derivatives include but are not limited to camptothecin, 10-hydroxycamptothecin, 9- aminocamptothecin, irinotecan, SN-38, edotecarin, topotecan and combinations thereof.
• Other antitumor agents include mitoxantrone, l-asparaginase, procarbazine, dacarbazine, hydroxycarbamide, pentostatin, tretinoin and combinations thereof.
• CTLA4 cytotoxic lymphocyte antigen 4
• CTLA4 compounds disclosed in U.S. Patent 6,682,736
• anti-proliferative agents such as other farnesyl protein transferase inhibitors, for example the farnesyl protein transferase inhibitors.
• CTLA4 antibodies that can be used in combination with compounds of the present invention include those disclosed in U.S. Patents 6,682,736 and 6,682,736 both of which are herein incorporated by reference in their entirety.
• Gene therapy agents may also be employed as anti-tumor agents such as TNFerade (GeneVec), which express TNFalpha in response to radiotherapy.
• TNFerade GeneVec
• statins may be used in combination with a compound of the present invention and pharmaceutical compositions thereof.
• Statins HMG-CoA reducatase inhibitors
• Atorvastatin LipitorTM, Pfizer Inc.
• Provastatin Provastatin
• Lovastatin MevacorTM, Merck Inc.
• Simvastatin ZaocorTM, Merck Inc.
• Fluvastatin LescolTM, Novartis
• Cerivastatin BoycolTM, Bayer
• Rosuvastatin CrestorTM, AstraZeneca
• Lovostatin and Niacin Niacin
• statin is selected from the group consisting of Atovorstatin and Lovastatin, derivatives and combinations thereof.
• Other agents useful as anti-tumor agents include Caduet.
• Ketone 3 can be further transformed to pyrazoles represented by formula 5 through a two step sequence involving enamine formation using N,N-dimethylformamide dimethyl acetal (DMF/DMA) followed by reaction with hydrazine.
• DMF/DMA N,N-dimethylformamide dimethyl acetal
• Introduction of the R 3 group on the pyrazole nitrogen can be accomplished through N-alkylation of a suitable alkyl halide (R 3 X), using a suitable base such as K 2 CO 3 , NaOH, DBU, NaH, LiHMDS, or Cs 2 CO 3 .
• pyrazoles During the alkylation reaction, two regioisomeric pyrazoles are usually formed. Pyrazoles represented by 6 can be separated and purified by silica gel chromatography or preparative HPLC and identified using NMR experiments including nuclear Overhauser effect spectroscopy.
• the protecting group can be methyl, trityl, benzyl, p-methoxybenzyl, tetrahydropyranyl, or cyclopropylmethyl.
• Various methods for phenol deprotection can be employed. These methods are known to those skilled in the art (e.g. see T. Greene and P. Wuts, "Protective Groups in Organic Synthesis", 3 rd Edition 1999, John Wiley & Sons).
• Suitable leaving groups include groups such as bromo, chloro, fluoro and methylsulfonyl.
• displacement of the leaving group can be accomplished using primary or secondary amines in a suitable solvent such as THF, ethanol, 2-propanol, toluene or xylenes at elevated temperatures to afford 2-amino heterocycles represented by 9.
• the protecting group can be methyl, trityl, benzyl, p-methoxybenzyl, tetrahydropyranyl, or methycyclopropane.
• Various methods for phenol deprotection can be employed and are known to those skilled in the art (e.g. see T. Greene and P. Wuts, "Protective Groups in Organic Synthesis", 3 rd Edition 1999, John Wiley & Sons).
• compounds of the general structure represented by 16 and 17 are prepared according to Method C.
• Preparation of isoxazole 11 is accomplished via the Vilsmeier reaction on 4-methylpyrimidine or picoline followed by condensation with hydroxylamine. Ring-opening of the isoxazole with aqueous sodium hydroxide is followed by condensation with hydrazine to afford aminopyrazole 13.
• a Sandmeyer reaction converts the amino group to an iodopyrazole (14) and, prior to the Suzuki coupling reaction, the pyrazole NH is either alkylated with a preferred R 3 group or protected with tetrahydropyranyl (THP) group to afford 15.
• THP tetrahydropyranyl
• a Suzuki coupling reaction of 15 with a boronic acid or boronic ester completes the synthesis of compounds represented by 16.
• the THP protecting group is used, it is removed under acidic conditions to afford 17.
• the protecting group can be methyl, trityl, benzyl, p-methoxybenzyl, tetrahydropyranyl, or cyclopropylmethyl.
• Various methods for phenol deprotection can be employed as discussed previously.
• compounds of the general structure represented by 20 and 22 are prepared according to Method D.
• Reaction of ketone 3 with carbon disulfide and dibromomethane in the presence of a suitable base such as Na 2 CO 3 , K 2 CO 3 or Cs 2 CO 3 in a solvent such as DMF or acetone affords dithietane 18.
• Reaction of 18 with amines at elevated temperatures in a suitable solvent such as toluene or xylenes affords vinylogous amide 19.
• Reaction of the thiol with an alkyl halide such as methyl iodide under basic conditions is followed by pyrazole formation using either hydrazine or substituted hydrazines to afford 5-aminopyrazoles such as 20.
• Intermediate 18 can also be reacted with sodium or potassium alkoxides to afford vinylogous ester 21 and these intermediates undergo the same sequence to afford 5-alkoxypyrazoles such as 22.
• the protecting group can be methyl, trityl, benzyl, p-methoxybenzyl, tetrahydropyranyl, or cyclopropylmethyl.
• Various methods for phenol deprotection can be employed as discussed previously.
• Ketone 23 can be transformed to 3,4,5-trisubstituted pyrazoles by tosylhydrazone formation followed by N-acylation of the tosylhydrazone with acid chlorides to form intermediates such as 25.
• the addition of 6 N HCI to 25 followed by gentle warming initiates a cyclization reaction and tosyl hydrazone elimination to form pyrazole 26.
• R 4 group on the pyrazole nitrogen can be accomplished through N-alkylation of a suitable alkyl halide (R 3 X), using a suitable base such as K 2 CO 3 , NaOH, DBU, NaH or Cs 2 CO 3 .
• a suitable base such as K 2 CO 3 , NaOH, DBU, NaH or Cs 2 CO 3 .
• the protecting group can be methyl, trityl, benzyl, p-methoxybenzyl, tetrahydropyranyl, or cyclopropylmethyl.
• Various methods for phenol deprotection can be employed as described previously.
• Examples 1 to 25 provide detailed synthetic steps for preparing several specific compounds of the present invention.
• Table 1 shows additional compounds that were prepared as Examples 26 to 30, and 32 to 127 using the methods described herein.
• Table 2 shows the biochemical and cellular data for the compounds of Examples 1 30, and 32 to 127. It is to be understood that the scope of the present invention is not limited in any way by the scope of the following examples and preparations.
• Et2 ⁇ diethyl ether
• DMF ⁇ /, ⁇ /-dimethylformamide
• THF tetrahydrofuran
• DCM diichloro-methane
• DMA dimethyl acetal
• DBU dimethyl acetal
• LiHMDS or LHMDS lithium hexamethyldisilazide
• TBME ferf-butyl methyl ether
• LDA Lithium Diisopropylamide
• DMSO dimethylsulfoxide
• MeOH methanol
• EtOH ethanol
• EtOAc ethyl acetate
• THF tetrahydrofuran
• Ac acetyl
• Me methyl
• Et (ethyl); and Ph phenyl
• the above compound was prepared as follows according to Method A. To an 8% (v:v) solution of trifluoroacetic acid in dichloromethane (10 mL) was added 4-[-3-(3-chloro-5-trityloxyphenyl)-1-isopropyl- 1H-pyrazol-4-yl]pyrimidine (242 mg, 0.43 mmol) and triethylsilane (80 ⁇ L, 58 mg, 0.5 mmol). The mixture was stirred at ambient temperature for 15 hours, then cautiously poured into saturated aqueous sodium hydrogen carbonate (40 mL). After stirring for an additional 45 minutes, the layers were separated and the aqueous phase extracted with dichloromethane (2 x 25 mL).
• the above compound was prepared as follows according to Method A. Following the procedure described in Example 2, using 4-[-3-(3-chloro-5-trityloxyphenyl)-1-ethyl-1 H-pyrazol-4-yl]pyrimidine in place of 4-[-3-(3-chloro-5-trityloxyphenyl)-1-isopropyl-1 H-pyrazol-4-yl]pyrimidine at ambient temperature, the title compound was obtained in 53% yield.
• the above compound was prepared as follows according to Method A. Following the procedure described in Example 2, using 4-(3-(3-chloro-4-methyl-5-(trityloxy)phenyl)-1-(pyridin-2-yl)-1 H-pyrazol-4- yl)pyrimidine in place of 4-[-3-(3-chloro-5-trityloxyphenyl)-1-isopropyl-1 H-pyrazol-4-yl]pyrimidine at ambient temperature, the title compound was obtained in 58% yield.
• the above compound was prepared as follows according to Method A. Following the procedure described in Example 2, using 2-(3-(3-chloro-4-methyl-5-(trityloxy)phenyl)-4-(pyrimidin-4-yl)-1H-pyrazol-1- yl)propanenitrile in place of 4-[-3-(3-chloro-5-trityloxyphenyl)-1-isopropyl-1 H-pyrazol-4-yl]pyrimidine at ambient temperature, the title compound was obtained in 37% yield.
• the above compound was prepared as follows according to Method A. To a mixture of 4-[3-(3- chloro-5-methoxyphenyl)-1H-pyrazol-4-yl]pyrimidine, (0.31 g, 1.1 mmol), prepared in the same way as in Example 1 except using 4-methylpyrimidine in place of picoline, and cesium carbonate (1.1 g, 3.2 mmol) in N,N-dimethylformamide (11 mL) was added tert-butyl 4-hydroxypiperidine-1-carboxylate (0.30 g, 1.1 mmol). The mixture was stirred at 70 0 C for 16 hours then concentrated to dryness to furnish 0.50 grams of an oil which was used in the next step without any purification.
• Method A To a mixture of 4-[3-(3- chloro-5-methoxyphenyl)-1H-pyrazol-4-yl]pyrimidine, (0.31 g, 1.1 mmol), prepared in the same way as in Example 1 except using 4-methyl
• the above compound was prepared as follows according to Method A. To a mixture of 4-[3-(3- chloro-5-methoxyphenyl)-1 H-pyrazol-4-yl]pyrimidine (1.0 g, 3.5 mmol) in N,N-dimethylformamide (23 mL) was added lithium bis(trimethylsilyl)amide (3.8 mL, 3.8 mmol) (1 M solution in tetrahydrofuran) dropwise. After stirring for 20 minutes, 1-chloro-3-iodopropane (1.1 mL, 11 mmol) was added and the mixture was allowed to stir for 16 hours.
• Method A To a mixture of 4-[3-(3- chloro-5-methoxyphenyl)-1 H-pyrazol-4-yl]pyrimidine (1.0 g, 3.5 mmol) in N,N-dimethylformamide (23 mL) was added lithium bis(trimethylsilyl)amide (3.8 mL, 3.8 mmol) (1 M solution
• the residue was treated with a 10% methanol in dichlororhethane solution, solids filtered and rinsed with the 10% methanol in dichloromethane solution, and filtrate concentrated to dryness.
• the residue was purified by high performance liquid chromatography to provide the title compound (0.010 g, 5%).
• the above compound was prepared as follows according to Method B. Boron tribromide (0.52 g. 2.1 mmol) was added to a solution of 4-[-3-(3-chloro-5-methoxyphenyl)-1-methyl-1H-pyrazol-4-yl]-2-(N- phenylamino)pyrimidine (249 mg, 0.64 mmol) in dichloromethane (20 mL) at 0 0 C. The cooling bath was removed and the resulting mixture was stirred at ambient temperature for 16 hours. The reaction was quenched upon addition of an ice/water mixture (20 mL) and ethyl acetate (20 mL).
• Aqueous methylamine (40% w / w , 0.57 mL, 6.6 mmol) was added to a solution of 4-[3-(3-chloro-5- methoxy-phenyl)-1-methyl-1 H-pyrazol-4-yl]-2-methanesulfonyl-pyrimidine (as obtained in Preparation b-1- b) (250 mg, 0.661 mmol) in dioxane (5 mL). The reaction mixture was stirred in a high pressure vessel at 85 0 C overnight, allowed to cool to room temperature, and concentrated in vacuo.
• the above compound was prepared as follows according to Method B.
• a mixture of 2-(3-(3- chloro-5-hydroxyphenyl)-4-(2-(methylsulfonyl)pyrimidin-4-yl)-1W-pyrazol-1-yl)acetonitrile (2.00 g, 5.14 mmol) and (S)-(+)-1-aminopropan-2-ol (3.86 g, 51.4 mmol) in anhydrous THF (50 mL) was heated to 80 0 C in an oil bath for 1.5 hours.
• the reaction mixture was diluted with ethyl acetate and methanol until homogeneous and was washed with sodium citrate buffer (1M, pH 4.5), and saturated sodium chloride (2 x 50 mL).
• the above compound was prepared as follows according to Method B. A mixture of 2-(3-(3- chloro-5-hydroxyphenyl)-4-(2-(methylsulfonyl)pyrimidin-4-yl)-1W-pyrazol-1-yl)acetonitrile (200 mg, 0.510 mmol) and 2-aminoethanol (0.31 mL, 5.1 mmol) were reacted at 80 0 C according to Example 11. After a similar work up, the crude residue was purified by reverse phase HPLC to provide the title compound (91 mg, 48%).
• the above compound was prepared as follows according to Method B. To a mixture of 3-chloro- 5- ⁇ 4-[2-(isobutylamino)pyrimidin-4-yl]-1-piperidin-4-yl-1 H-pyrazol-3-yl ⁇ phenol (0.15 g, 0.32 mmol) in acetonitrile (4 mL) was added a 51 % aqueous formaldehyde solution (75 mL, 0.95 mmol). The mixture was allowed to stir for 20 minutes then sodium cyanoborohydride (60 mg, 0.95 mmol) was added and the mixture was allowed to stir for 10 minutes. Acetic acid was added dropwise until neutral and the entire mixture was concentrated to dryness.
• Method B To a mixture of 3-chloro- 5- ⁇ 4-[2-(isobutylamino)pyrimidin-4-yl]-1-piperidin-4-yl-1 H-pyrazol-3-yl ⁇ phenol (0.15 g, 0.32 mmol) in acetonit
• the above compound was prepared as follows according to Method B. Following the procedure described in Example 17, using 3-chloro-5-(4- ⁇ 2-[(2-hydroxy-2-methylpropyl)amino]pyrimidin-4-yl ⁇ -1- piperidin-4-yl-1/-/-pyrazol-3-yl)phenol in place of 3-chloro-5- ⁇ 4-[2-(isobutylamino)pyrimidin-4-yl]-1-piperidin- 4-yl-1 H-pyrazol-3-yl ⁇ phenol, the title compound was obtained in 57% yield.
• the above compound was prepared as follows according to Method C. Following the procedure described in Example 20, using 2,3-dimethyl-5-(4-(pyrimidin-4-yl)-1H-pyrazol-3-yl)phenol in place of 3-(3- (3-methoxy-2,5-dimethylphenyl)-4-(pyrimidin-4-yl)-1H-pyrazol-1-yl)propanenitrile, the title compound was obtained in 46% yield.
• the above compound was prepared as follows according to Method C. Boron tribromide (0.084 ml_ of 1.0 M solution in DCM, 0.84 mmol) was added to a solution of 4-(3-(5-fluoro-2-methoxyphenyl)-1- methyl-1W-pyrazol-4-yl)pyrimidine (80 mg, 0.28 mmol) in dichloromethane (2 mL) at 0 0 C. The resulting suspension was stirred at 0 °C for 45 minutes. The reaction was diluted with dichloromethane (10 mL) and quenched by the slow addition of saturated aqueous sodium hydrogen carbonate (10 mL). The resulting mixture was stirred at ambient temperature until two phases were evident.
• the nitrogen bubbler was removed shortly after the addition and the mixture was sealed and placed in a Biotage InitiatorTM Microwave Synthesizer and heated at 130 0 C for 1 hour. After cooling to ambient temperature, the crude reaction mixture was partitioned between saturated aqueous sodium chloride (100 mL) and ethyl acetate (50 mL). The layers were separated and the aqueous phase extracted with ethyl acetate (2 x 50 mL). The combined organic extracts were washed with water (2x50 mL), brine (50 mL), dried (anhydrous magnesium sulfate), filtered and concentrated to dryness.
• aqueous sodium chloride 100 mL
• ethyl acetate 50 mL
• the combined organic extracts were washed with water (2x50 mL), brine (50 mL), dried (anhydrous magnesium sulfate), filtered and concentrated to dryness.
• the pyrazole regiochemistry of the major regioisomer was confirmed as drawn above by running a nuclear Overhauser effect (NOE) spectroscopy experiment in C 6 D 6 prior to trityl deprotection.
• the protected phenol (90.0 mg, 0.147 mmol) was dissolved in DCM (3 mL) and treated with Et 3 SiH (100 ⁇ L) followed by TFA (0.5 mL). After 30 minutes, LC/MS analysis indicated complete deprotection and showed the two regioisomers resolved by LCMS. After removing the solvents, the product was purified via silica gel chromatography, eluting with 60/40, 70/30 then 80/20 EtOAc / Pet Ether.
• the aqueous layer was further extracted with a 1 :1 mixture of Et 2 O / EtOAc (3 x 75 mL). The combined organic extracts were washed with water (3 x 75 ml_) and brine (1 x 100 mL) then dried over MgSO 4 . The solvents were removed to afford an orange foam (7.3 g, 85%) used in the next step without further purification.
• the above compound was prepared as follows according to Method D. To a solution of 1-(3- methyl-5-(trityloxy)phenyl)-2-(pyrimidin-4-yl)ethanone (8 g, 17.0 mmol) in ⁇ /, ⁇ /-dimethylformamide (86 mL) was added cesium carbonate (11.1 g, 2 equiv), followed by carbon disulfide (3.08 mL, 3 equiv) and dibromomethane (3.54 mL, 3 equiv). The mixture was stirred at ambient temperature for 6 hours. The solution was poured into ice water and extracted with a mixture of 50% ethyl acetate and 50% diethyl ether (4x).
• the pyrazole formed after the two step sequence involving enamine formation using DMF/DMA followed by reaction with hydrazine was alkylated with (2- bromoethoxy)(tert-butyl)dimethylsilane. Both the tetrahyropyran and tertbutyl dimethyl silyl protecting groups are removed in the subsequent oxidation step. Displacement of the sulfone with amines according to Method B gave the final targets.
• Example 136 was prepared following the procedure described in Example 13 (see also general method B) except 2-(3-(3-hydroxy-4,5-dimethylphenyl)-4-(2-(methylsulfonyl)pyrimidin-4-yl)-1 H-pyrazol-1 - yl)acetonitrile was used in place of 3-(3-(3-hydroxy-5-methylphenyl)-4-(2-(methylsulfonyl)-pyrimidin-4-yl)- 1H-pyrazol-yl)propanenitrile and the alkylating step was carried out with bromoacetonitrile.
• Example 139 2-(3-(3-Flouro-5-hydroxyphenyl)-4-(2-((S)-2-hydroxypropylamino)pyrimidin-4-yl)-1 H-pyrazol-1- yl)acetonitrile (Example 139) was prepared according to method C except starting with 4-methyl-2- (methylthio)pyrimidine gives 4-(3-iodo-1-(tetrahydro-2H-pyran-2-yl)-1 H-pyrazoi-4-yl)-2-
• Example b-8-a The title compound was prepared following the procedure described in Example b-8-a except 4- (3-(3-fluoro-5-methoxyphenyl)-1H-pyrazol-4-yl)-2-(methylthio)pyrimidine was used in place of tert-butyl 3- (3-(3-methoxy-5-methylphenyl)- 4-(2-(methylthio)pyrimidin-4-yl)-1/-/-pyrazol-1-yl)azetidine-1 carboxylate to give 4-(3-(3-fluoro-5-methoxyphenyl)-1H-pyrazol-4-yl)-2-(methylsulfonyl)pyrimidine in quantitative yield.
• Example c-1-c he title compound was prepared following the procedure described in Example c-1-c except 3- fluoro-5-methoxyphenylboronic acid was used in place of 3-methoxy-2,5-dimethylphenylboronic acid and 4-(3-iodo-1-tetrahydro-2/-/-pyran-2-yl)-1H-pyrazol-4-yl)-2-(methylthio)pyrimidine was used in place of 4-(3- iodo-1-tetrahydro-2W-pyran-2-yl)-1H-pyrazol-4-yl)pyrimidine.
• the mixture was heated in a microwave at 13O 0 C for 1 hour rather than in an oil bath at 100 0 C for 10 hours.
• the supernatant solution was decanted into a separate flask.
• the solids were rinsed with a small amount of DMF and the rinse combined with the supernatant solution.
• the solvent was removed en vacuo and the residue was taken up in methanol and treated with HCI (4N in dioxane, 4 eq).
• the mixture was stirred at ambient temperature for 15 minutes then reduced to minimum volume.
• the residue was triturated with dichloromethane/ te/t-methyl butyl ether ( ⁇ 1 :1) and the solids collected by filtration to yield clean product.
• Example 140 3-(1-(cyanomethyl)-4-(2-(2-hydroxyethylamino)primidin-4-yl)-1H-pyrazole-3-yl)-5- hydroxybenzonitrile (Example 140) was prepared according to method B and subsequent conversion of the chloro to the cyano following the procedure of Leadbeater and Arvela; JOC 2003, 68, 9122-9125.
• Example b-6-a The title compound was prepared following the procedure described in Example b-6-a except 4- (3-(3-chloro-4-fluoro-5-methoxyphenyl)-1 H-pyrazol-4-yl)-2-(methylthio)pyrimidine was used in place of 3- (3-(3-methyl-5-(trityloxy)phenyl)-4-(2-(methylthio)pyrimidin-4-yl)-1/-/-pyrazol-yl)propanenitrile to give 4-(3- (3-chloro-4-fluoro-5-methoxyphenyl)-1 H-pyrazol-4-yl)-2-(methylsulfonyl)pyrimidine in 73% yield.
• Example 13 2-(3-(3-chloro-4-fluoro-5- hydroxyphenyl)-4-(2-(2-hydroxyethylamino)pyrimidin-4-yl)-1 H-pyrazol-1-yl)acetonitrile, starting from methyl 4-fluoro-3-methoxy-5-methylbenzoate and 4-methyl-2-(methylthio)pyrimidine.
• Raf kinase activity is measured in vitro by determining transfer of radiolabeled 32-P phosphate from ATP to the specific Raf substrate Mek1.
• Full-length wild type b-Raf is expressed in recombinant form and purified from bacterial or insect cells.
• Recombinant Mek1 is purified from E. coli bacterial cells.
• the full-length wild type Mek1 is used as the b-Raf substrate.
• G2 the full-length K97R Mek1 mutant is used as the b- Raf substrate.
• In vitro kinase assays are performed in solution containing the following; 50 mM Hepes (pH 7.4), 5 nM b-Raf, 0.8 ⁇ M Mek1 , 10 mM MgCI2, 25 ⁇ M ATP, 0.002% (v/v) Tween-20, 5 ⁇ g/mL leupeptin, 1.2 mM DTT, 2% (v/v) DMSO, 0.2-1.0 ⁇ Ci [ ⁇ - 32 P]ATP per well.
• the assays are performed in wells of a 96 well polypropylene round bottom plate, each well containing 43.5 ⁇ l_ assay mix and 1.5 ⁇ l_ inhibitor compound or DMSO vehicle. 15 ⁇ L of b-Raf mix is added and the plate is shaken on a plate shaker and preincubated for 10 minutes at ambient temperature.
• the reaction is started by addition of 15 ⁇ L ATP mix and shaking. The reaction is terminated after 40 minutes by addition of 25 ⁇ L 0.5 M EDTA (pH 7.4).
• 60 ⁇ L of the stopped reaction is transferred to a well of a 96-well nylon 66 Biodyne A membrane Silent Screen filter plate (Nalge/Nunc: 256081).
• the wells are filtered and washed five times with 0.85% phosphoric acid.
• the filter is placed in a tray with about 50 mL 0.85% phosphoric acid and gently rotated for 10 minutes on an orbital shaker. The procedure is repeated once with fresh 0.85% phosphoric acid.
• Raf kinase activity is calculated from the specific activity of [32-P] ATP, 32-P incorporation into Mek1 , and the concentration of b-Raf.
• Healthy growing human melanoma A2058 cells (harboring a b-Raf mutation) are used for the assay.
• A2058 cells are grown in 10% FBS DMEM medium. When the cells are near 85%+ confluence, the cells are rinsed with PBS once and trypsinized with trypsin/EDTA for 3 minutes. The cells are resuspended in 10% FBS DMEM and are centrifuged down at 1000 rpm for 5 minutes. The cells are resuspended in 0.5% FBS DMEM and counted on a cell counter. The cells are seeded at 50,000 cells/well in a volume of 100 ⁇ L/well in 0.5% FBS DMEM in a 96 well flat-bottom plate. The negative control wells receive only 100 ⁇ l_ of 0.5% FBS DMEM medium without cells. The plate is incubated overnight in a cell culture incubator with 5% CO 2 at 37 0 C.
• testing compounds are prepared in 0.5% FBS DMEM medium and serially diluted at 1:3 for 11 test concentrations. Each concentration of the compounds is tested in duplicate.
• the compound solutions are added at 25 ⁇ L/well to the corresponding wells in the cell plate, and 25 ⁇ L/well of the vehicle (0.5% DMSO in 0.5% FBS DMEM) is added to the negative control wells (no cells) and the positive control wells (cells without compounds).
• the plate is incubated for 1 hour in a cell culture incubator with 5% CO 2 at 37 °C. After 1 hour of incubation, the medium is removed, 100 ⁇ L/well of cell lysis buffer is added into the cell plate, and the plate is shaken for 15 minutes at room temperature.
• the cell lysates are transferred to an ELISA plate (pre-coated with anti-Mek1 anti-body, Cell Signaling #2352), and the plate is incubated with gentle shaking for 2 hours at room temperature. After 2 hours, the contents of the wells are aspirated and the wells are washed 4 times with wash buffer. 100 ⁇ L of phospho-Mek1/2 detection antibody (Cell Signaling #9121) is added into each well and the plate is incubated with gentle shaking for 1 hour at room temperature. After 1 hour, the wells are aspirated and washed 4 times with wash buffer.
• phospho-Mek1/2 detection antibody Cell Signaling #9121

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